α-amylase variants

Abstract

The present invention relates to α-amylase variants of a parent α-amylase that have improved cleaning performance when compared to the parent α-amylase. The invention also relates to polynucleotides encoding the variants, nucleic acid constructs, vectors, and host cells comprising the polynucleotides, and methods of introducing the variants of the invention.

Description

[Technical Field]

[0001] Sequence listing reference This application includes a computer-readable sequence listing, which is incorporated herein by reference.

[0002] The present invention relates to α-amylase variants having α-amylase activity. The present invention further relates to a vector and host cell comprising a polynucleotide encoding such variant, a gene encoding such variant, and which may also enable the production of such variant. The present invention further relates to use in household cleaning and treatment, particularly in laundry and dishwashing, including hand washing and / or automatic washing, and / or hand washing and / or automatic dishwashing. The present invention is particularly useful for washing laundry. [Background technology]

[0003] α-Amylases (α-1,4-glucan-4-glucanohydrolase, EC3.2.1.1) comprise a group of enzymes that catalyze the hydrolysis of starch and other linear and branched 1,4-glucosidoligosaccharides and polysaccharides. They are well known for their long-standing use in household products, for example, the first bacterial α-amylase from B. licheniformis, also known as Termamyl, has been used. Alkaline amylases such as AA560 (International Publication No. 2000 / 060060) and SP707 (described in Tsukamoto et al., 1988, Biochem. Biophys. Res. Comm. 151; 25-31) are useful in detergents, and it is known that these and other amylases are modified to improve their stability in detergents. For example, to improve the stability of this amylase, International Publication No. 96 / 23873 describes deleting amino acids 181+182 or 183+184 (Sequence ID 7 in International Publication No. 96 / 23873) of SP707. International Publication No. 96 / 23873 further describes modifying SP707 amylase by substituting M202 with, for example, leucine, to stabilize the molecule against oxidation.

[0004] In particular, improving the performance of amylose-degrading enzymes in an environmentally friendly manner has always been a challenge. Furthermore, for environmental reasons, it is becoming increasingly important to provide amylose-degrading enzymes that are effective and preferably rapid in washing cycles, often using less water and / or lower temperatures. Therefore, it is desirable to have an amylose-degrading enzyme that exhibits improved properties, such as specific activity, compared to the parent polypeptide. The present invention provides a mutant polypeptide that has α-amylase activity and improved properties, such as the removal of starch-containing stains, or enzymatic washing advantages or specific activity, compared to the hybrid and / or parent polypeptide. Particularly preferred amylose-degrading enzymes function at low temperatures and / or in short washing cycles, while simultaneously maintaining or improving other desirable properties such as specific activity (amylose-degrading activity), stability, and / or washing performance.

[0005] Accordingly, an object of the present invention is to provide an α-amylase variant that exhibits improved properties, such as removal of starch-containing stains or specific activity, and can be used in washing, dishwashing, and / or washing processes at low temperatures such as 5 to 40°C, compared to the parent polypeptide. A further object of the present invention is to provide an α-amylase variant having improved washing performance compared to the parent α-amylase or compared to any of the α-amylases of SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14. [Overview of the project] [Means for solving the problem]

[0006] The present invention uses sequence number 1 for numbering purposes to represent 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, The present invention relates to an α-amylase variant of a parent α-amylase that includes modifications at one or more positions corresponding to positions 360, 361, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant is a parent polypeptide with at least 60%, at least 65%, at least 70%, at least 75%, and at least 8 0%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0007] The present invention uses sequence number 1 for numbering purposes to represent 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 1 74, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 361, 362, 365, 371, 383, 390, 394, 395, 405, 409 The present invention relates to an α-amylase variant of a parent α-amylase that includes modifications at one or more positions corresponding to positions 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant is at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, and at least 87% of the parent polypeptide. The variant has sequence homology of at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, provided that the variant has sequence homology of less than 100%, and the variant has improved washing performance compared to the parent polypeptide, which is preferably SEQ ID NO: 1, SEQ ID NO: 2, or SEQ ID NO: 3.

[0008] The present invention also relates to a polynucleotide encoding an α-amylase variant according to the present invention, a nucleic acid construct comprising a polynucleotide encoding a variant according to the present invention, an expression vector comprising a polynucleotide encoding a variant according to the present invention, or a host cell comprising a polynucleotide encoding a variant according to the present invention.

[0009] The present invention also relates to a method for producing an α-amylase mutant, comprising (a) culturing host cells of the present invention under conditions suitable for mutant expression, and (b) recovering the mutant.

[0010] The present invention further uses Sequence ID No. 1 for numbering: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 13 6, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 24 4, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 361, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 45 The present invention relates to a method for obtaining an α-amylase mutant, comprising introducing a modification at one or more positions corresponding to positions 5, 457, 458, 470, 472, 473, 474, 475, and 476 into a parent α-amylase, wherein each modification is independently a substitution or deletion, and the mutant has α-amylase activity, and comprising the steps of introduction and recovery of the mutant.

[0011] definition In embodiments of this invention, the following definitions apply. Note that unless otherwise explicitly indicated in the context, the singular forms "a," "an," and "the" include multiple subjects.

[0012] In this specification, any reference to a value or parameter "about" includes aspects relating to that value or parameter itself. For example, a description referring to "about X" includes aspect "X".

[0013] Throughout this specification, values ​​expressed in range form should be interpreted flexibly to include not only the numerical values ​​explicitly listed as limits to the range, but also all individual numerical values ​​or subranges contained within the range, as if each numerical value and subrange were explicitly listed. For example, the range "approximately 0.1 percent to approximately 5 percent" or "approximately 0.1 percent to approximately 5 percent" should be interpreted to include not only approximately 0.1 percent to approximately 5 percent, but also the individual values ​​within the indicated range (e.g., 1 percent, 2 percent, 3 percent, and 4 percent), and subranges (e.g., 0.1 percent to 0.5 percent, 1.1 percent to 2.2 percent, 3.3 percent to 4.4 percent). Unless otherwise indicated, the description "approximately X to Y" is equivalent to "approximately X to approximately Y". Similarly, unless otherwise indicated, the description "approximately X, Y, or approximately Z" is equivalent to "approximately X, approximately Y, or approximately Z".

[0014] Unless otherwise defined or explicitly indicated by the context, all technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art to which this invention pertains.

[0015] Domains A, B, and C: The structure of α - amylase includes three distinct domains, A, B, and C. See, for example, Machius et al., 1995, J. Mol. Biol. 246; 545 - 559. The term "domain" means a region of a polypeptide that forms a distinct and independent structure of the whole molecule. α - amylase consists of an active site designated as domain A, a somewhat long loop between β - sheet 3 and α - helix 3 designated as domain B, and a β / α - 8 barrel designated as domain C, and may also have a carbohydrate - binding domain in some cases (for example, WO 2005 / 001064 pamphlet; Machius et al., supra).

[0016] The domains of α - amylase can be determined, for example, by structural analysis using crystallographic techniques. An alternative method for determining the domains of α - amylase is to align the amino acid sequence of α - amylase with that of another α - amylase for which the domains have been determined. For example, the sequence that aligns with the B - domain sequence of an α - amylase for which the B - domain has been determined can be regarded as the B - domain of a given α - amylase.

[0017] Allelic variants: The term "allelic variant" means any of two or more alternative forms of a gene that occupy the same chromosomal locus. Mutations can naturally generate allelic variations of a gene, resulting in genetic polymorphisms within a population. The gene mutation may be silent (no mutation within the encoded polypeptide) or may encode a polypeptide with an altered amino acid sequence. An allelic variant of a polypeptide is a polypeptide encoded by an allelic variant of the gene.

[0018] α-Amylase: The term "α-amylase" is synonymous with "polypeptide having α-amylase activity". "α-Amylase activity" means the activity of α-1,4-glucan-4-glucanohydrolase, E.C. 3.2.1.1, which is composed of a group of enzymes that catalyze the hydrolysis of starch and other linear and branched 1,4-glucosidic oligosaccharides and polysaccharides. Thus, as used herein, the term "α-amylase" refers to an enzyme having α-amylase activity (enzyme classification: EC 3.2.1.1) that hydrolyzes the α-linkages of large α-linked polysaccharides such as starch and glycogen to yield glucose and maltose. The terms "α-amylase" and "amylase" may be used interchangeably and have the same meaning and purpose herein. For the purposes of the present invention, α-amylase activity is determined according to the procedure described under "Method" in the subheading below. In one aspect, the α-amylase of the present invention has an amino acid sequence with at least 20%, for example at least 40%, at least 50%, at least 60%, for example at least 65%, for example at least 70%, for example at least 75%, for example at least 80%, for example about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, but less than 100% sequence homology to the polypeptide of SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 or 14.

[0019] Amino Acids: As used herein, the term “amino acids” refers to the 20 standard genetically encoded amino acids and their corresponding stereoisomers in “d” forms (compared to the natural “l” forms), omega amino acids, other naturally occurring amino acids, unconventional amino acids (e.g., α,α-disubstituted amino acids, N-alkyl amino acids, etc.), and chemically derivatized amino acids. Chemical derivatization of one or more amino acids may be achieved by reacting them with functional side groups. Examples of such derivatized molecules include molecules in which a free amino group is derivatized to form an amine hydrochloride, a p-toluenesulfonyl group, a carboxybenzoxy group, a t-butyloxycarbonyl group, a chloroacetyl group, or a formyl group. A free carboxyl group may be derivatized to form salts, methyl and ethyl esters or other types of esters and hydrazides. A free hydroxyl group may be derivatized to form O-acyl or O-alkyl derivatives. Examples of chemical derivatives include peptides, which contain naturally occurring amino acid derivatives of the 20 standard amino acids. For example, 4-hydroxyproline may be substituted with proline, 5-hydroxylysine may be substituted with lysine, 3-methylhistidine may be substituted with histidine, and homoserine may be substituted with serine and ornithine for lysine. Derivatives may also include peptides containing one or more additions or deletions, as long as the required activity is maintained. Other modifications include terminal modifications such as amidation, amino-terminus acylation (e.g., acetylation or thioglycolic acid amidation), and terminal carboxyamidation (e.g., by ammonia or methylamine).

[0020] Where amino acids are specifically listed as "alanine," "Ala," or "A," unless otherwise explicitly stated, this term refers to both l-alanine and d-alanine. Other unconventional amino acids may also be suitable components of the polypeptide of the present invention, insofar as the polypeptide retains the desired functional properties. With respect to the peptides shown, each encoded amino acid residue is represented, if necessary, by a single letter corresponding to the common name of a conventional amino acid. In one embodiment, the polypeptide of the present invention contains or consists of an l-amino acid.

[0021] Catalytic domain: The term "catalytic domain" refers to the region of an enzyme that contains the catalytic mechanism.

[0022] cDNA: The term "cDNA" refers to DNA molecules that can be prepared by reverse transcription from mature, spliced ​​mRNA molecules obtained from eukaryotic cells. cDNA lacks intron sequences that may be present in the corresponding genomic DNA. The initial primary RNA transcript is a precursor of mRNA that has been processed through a series of steps, including splicing, before the mature, spliced ​​mRNA appears.

[0023] Coding sequence: The term "coding sequence" refers to a polynucleotide that directly identifies the amino acid sequence of a variant. The boundaries of a coding sequence are generally determined by an open reading frame that typically begins with a start codon such as ATG, GTG, or TTG and ends with a stop codon such as TAA, TAG, or TGA. A coding sequence may be DNA, cDNA, or a synthetic or recombinant polynucleotide.

[0024] Regulatory Sequence: The term "regulatory sequence" means a nucleic acid sequence required for the expression of the polynucleotide encoding the variant of the present invention. Each regulatory sequence may be native (i.e., derived from the same gene) or exogenous (i.e., derived from a different gene) to the polynucleotide encoding the variant, or may be native or exogenous to each other. Examples of such regulatory sequences include, but are not limited to, leaders, polyadenylation sequences, propeptide sequences, promoters, signal peptide sequences, and transcriptional terminators. At a minimum, a regulatory sequence includes a promoter and transcriptional and translational stop signals. The regulatory sequence may be provided with a linker for introducing a specific restriction site to facilitate linking of the coding region of the polynucleotide encoding the variant to the regulatory sequence.

[0025] Corresponding: As used herein, the term "corresponding" refers to a method for determining the specific amino acids of a sequence that references a specific amino acid sequence. For example, when a specific amino acid position is referenced for the purposes of the present invention, a person skilled in the art can align the referenced amino acid sequence with the other amino acid sequence to determine which specific amino acids may be the target in the other amino acid sequence. Aligning the other amino acid sequence with, for example, the sequences described in SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14, or any other sequences listed herein, is described elsewhere in this specification. Alternative alignment methods may be used, which are known to a person skilled in the art.

[0026] Dishwashing composition: The term “dishwashing composition” as used herein refers to any form of composition for washing dishes. A dishwashing composition may be a liquid dishwashing composition or a powder dishwashing composition, and optionally, the composition may be in the form of a unit dose. A unit dose of dishwashing composition may contain two or more compartments, the compartments of which may contain powder, liquid, or a combination thereof.

[0027] Benefits of Enzymatic Cleaning Power: As used herein, the term “benefits of enzymatic cleaning power” refers to the advantageous effects that enzymes may impart to a detergent compared to the same detergent without enzymes. Key cleaning benefits that enzymes may provide include stain removal that is completely or minimally visible after washing and / or washing, preventing or reducing the reattachment of loosened dirt during the washing process (also known as anti-reattachment effect), and completely or partially restoring the whiteness of fabrics that were originally white but have become grayish or yellowish after repeated use and washing (also known as whitening effect). Benefits of fabric care may be important to the benefits of enzymatic cleaning power, even if they are not directly related to catalytic stain removal or prevention of dirt reattachment. Examples of the benefits of such fabric care include preventing or reducing dye transfer from one fabric to another or to a different part of the same fabric (an effect also known as dye transfer inhibition or back-dye prevention), reducing the tendency for pilling by removing protruding or broken fibers from the fabric surface, or removing existing pilling or fuzz (an effect also known as anti-pilling), improving the flexibility of the fabric, clarifying the color of the fabric, and removing particulate dirt trapped in the fibers of the fabric or garment. Enzyme bleaching generally offers the additional benefit of enzymatic cleaning power, where catalytic activity is used to catalyze the formation of bleaching components such as hydrogen peroxide or other peroxides.

[0028] Expression: The term "expression" includes any steps involved in the generation of a variant, including but not limited to transcription, post-transcriptional modification, translation, post-translational modification, and secretion.

[0029] Expression vector: The term "expression vector" refers to a linear or circular DNA molecule containing polynucleotides encoding a variant, which is operably ligated to further nucleotides that result in expression.

[0030] Fragment: As used herein, the term "fragment" refers to a polypeptide having one or more (e.g., several) amino acids without amino and / or carboxyl ends of any one of the parent sequences disclosed herein, e.g., SEQ ID NOs: 1-14, and the fragment has α-amylase activity. In one embodiment, the fragment comprises at least 200 consecutive amino acid residues of SEQ ID NOs: 1-14, e.g., at least 300 consecutive amino acid residues of SEQ ID NOs: 1-14, or at least 350 consecutive amino acid residues, or at least 400 consecutive amino acid residues, or at least 450 consecutive amino acid residues.

[0031] Fusion polypeptide: As used herein, the term "fusion polypeptide" refers to a polypeptide comprising amino acid sequences derived from two or more species, for example, two, three, or four species. Such fusion polypeptides may be produced by molecular techniques well known to those skilled in the art. The fusion polypeptides of the present invention have α-amylase activity. In particular, the fusion polypeptides of the present invention are composed, for example, of the A and C domains of one α-amylase and the B domain derived from another α-amylase. Hybrid polypeptide (or "hybrid") and fusion polypeptide can be used interchangeably and have the same meaning and purpose.

[0032] Hard surface cleaning: As used herein, the term “hard surface cleaning” refers to cleaning any hard surface, including floors, roofs, cars, kitchen or bathroom surfaces, and dishwashing (dishwashing).

[0033] Heterogenetic polynucleotides: The term “heterogenetic polynucleotide” is defined herein as a polynucleotide that is not native to a host cell, a native polynucleotide whose coding region has been structurally modified, a native polynucleotide whose expression has been quantitatively altered as a result of recombinant DNA technology, such as manipulating DNA with a different (foreign) promoter, or a native polynucleotide in a host cell that has one or more additional copies of a polynucleotide whose expression is quantitatively altered. A “heterogenetic gene” is a gene that contains heterogenetic polynucleotides.

[0034] Heterogeneous: The term “heterogeneous” in relation to a host cell means a polypeptide or nucleic acid that is not naturally present in the host cell. The term “heterogeneous” in relation to a polypeptide or nucleic acid means that a regulatory sequence, such as a promoter or a domain of the polypeptide or nucleic acid, does not naturally bind to the polypeptide or nucleic acid; that is, the regulatory sequence originates from a gene other than the gene encoding the variant.

[0035] High stringency conditions: The term "high stringency conditions" means that for probes of at least 100 nucleotides in length, pre-hybridize and hybridize at 42°C in 5×SSPE, 0.3% SDS, 200 micrograms / ml shear and denatured salmon sperm DNA, and 50% formamide, followed by a standard Southern blotting procedure for 12-24 hours. Finally, wash the carrier material three times each with 2×SSC and 0.2% SDS at 65°C for 15 minutes.

[0036] Host cell: The term "host cell" means any microbial or plant cell into which the nucleic acid construct or expression vector containing the polynucleotide of the present invention is introduced. Methods of introduction include, but are not limited to, protoplast fusion, translocation, transformation, electroporation, conjugation, and transduction. In some embodiments, the host cell is an isolated recombinant host cell partially or completely isolated from at least one other component, including but not limited to proteins, nucleic acids, or cells.

[0037] Hybridization: The term "hybridization" means substantially pairing complementary strands of nucleic acids using a standard Southern blotting procedure. Hybridization may be performed under medium, medium-high, high, or very high stringency conditions. Medium stringency conditions involve pre-hybridizing and hybridizing at 42°C for 12-24 hours in 5×SSPE, 0.3% SDS, 200 micrograms / ml of sheared and denatured salmon sperm DNA and 35% formamide, followed by washing three times at 55°C for 15 minutes each with 0.2×SSC and 0.2% SDS. For medium to high stringency conditions, pre-hybridize and hybridize in 5×SSPE, 0.3% SDS, 200 micrograms / ml of sheared and denatured salmon sperm DNA, and 35% formamide at 42°C for 12-24 hours, then wash three times each at 60°C with 0.2×SSC and 0.2% SDS for 15 minutes each. For high stringency conditions, pre-hybridize and hybridize in 5×SSPE, 0.3% SDS, 200 micrograms / ml of sheared and denatured salmon sperm DNA, and 50% formamide at 42°C for 12-24 hours, then wash three times each at 65°C with 0.2×SSC and 0.2% SDS for 15 minutes each. For very high stringency conditions, pre-hybridize and hybridize in 5×SSPE, 0.3% SDS, 200 micrograms / ml shear and denatured salmon sperm DNA, and 50% formamide at 42°C for 12-24 hours, then wash three times each at 70°C for 15 minutes using 0.2×SSC and 0.2% SDS.

[0038] Improved Properties: The term "improved properties" is defined herein as features associated with an improved variant compared to the parent α-amylase. Such improved properties include, but are not limited to, improved amylose decomposition activity, improved catalytic efficiency, improved catalytic reaction rate, improved chemical stability, improved oxidative stability, improved pH activity, improved pH stability, improved specific activity, improved substrate binding, improved substrate cleavage, improved substrate specificity, improved substrate stability, improved surface properties, improved thermal activity, improved thermal stability, improved cleaning performance such as stain removal performance, e.g., improved performance in removing starch-containing stains, stains, and preventing graying, stability, e.g., thermal stability, pH stability, or stability in the presence of a builder containing a chelating agent, stability in powder, liquid, or gel detergent formulations or dishwashing compositions, changing temperature-dependent performance, and activity profile, pH activity, substrate specificity, product specificity, and chemical stability.

[0039] Improved Cleaning Performance: The term "improved cleaning performance" is defined herein as an improvement in the cleaning performance of an amylase compared to the cleaning performance of the parent amylase, or compared to any of the α-amylases of SEQ ID NO: 1, SEQ ID NO: 2, or SEQ ID NO: 3, for example, by improving stain removal. Improved cleaning performance may be measured by comparing so-called intensity values. Improved cleaning performance is preferably measured according to the examples. Cleaning performance is improved if the improvement factor (IF) is greater than 1.0, preferably greater than 1.05, in one or more of the conditions listed in the examples. The cleaning conditions are described in the Examples section. The term "cleaning performance" generally includes cleaning hard surfaces, such as dishwashing, but also includes cleaning performance on textiles, such as laundry, and cleaning performance in industrial and commercial cleaning. Improved cleaning performance may be measured by comparing delta intensities as defined herein.

[0040] Isolated: The term “isolated” means a substance in a form or environment that does not exist in nature. Non-limiting examples of isolated substances include: (1) any substance that does not exist in nature; (2) any enzyme, mutant, nucleic acid, protein, peptide or cofactor from which one or more or all of the naturally occurring components to which it binds in nature have been partially removed; (3) any substance that has been modified by human intervention from a substance found in nature; or (4) any substance that has been modified by increasing the amount of the substance compared to other components that bind it naturally (e.g., multiple copies of the gene encoding the substance, or the use of a promoter stronger than the promoter that naturally binds to the gene encoding the substance). Isolated substances may be present in fermentation broth samples. In one embodiment, the present invention relates to an isolated α-amylase mutant.

[0041] Intensity Value: Cleaning performance is measured as luminance, which is the intensity of light reflected from the sample when illuminated with white light. When a sample is stained, the intensity of the reflected light is lower than that of a clean sample. Therefore, cleaning performance can be measured using the intensity of the reflected light, and in this case, a higher intensity value correlates with higher cleaning performance. Color measurement is performed by capturing an image of the cleaned fabric using a specialized flatbed scanner (Kodak iQsmart, Kodak). To extract the light intensity value from the scanned image, the 24-bit pixel values ​​from the image are converted to red, green, and blue (RGB) values. The intensity value (Int) is calculated by adding the RGB values ​​together as a vector and then obtaining the length of the resulting vector.

number

[0042] Low stringency conditions: The term "low stringency conditions" means that for probes of at least 100 nucleotides in length, pre-hybridize and hybridize at 42°C in 5×SSPE, 0.3% SDS, 200 micrograms / ml shear and denatured salmon sperm DNA, and 25% formamide, followed by a standard Southern blotting procedure for 12-24 hours. Finally, wash the carrier material three times each with 0.2×SSC and 0.2% SDS at 50°C for 15 minutes.

[0043] Low temperature: "Low temperature" refers to temperatures of 5-40°C, or 5-35°C, or 5-30°C, or even 5-25°C, or 5-20°C, or 5-15°C, or 5-10°C. Preferably, "low temperature" refers to temperatures of 10-35°C, preferably 10-30°C, or 10-25°C, or 10-20°C.

[0044] Mature polypeptide coding sequence: The term "mature polypeptide coding sequence" refers to a polynucleotide that codes for a mature polypeptide possessing α-amylase activity.

[0045] Mature polypeptide: The term “mature polypeptide” refers to a polypeptide in its final form after translation and any post-translational modifications, such as N-terminal processing, C-terminal truncation, glycosylation, phosphorylation, etc. It is known in the art that host cells can produce a mixture of two or more different mature polypeptides expressed by the same polynucleotide (i.e., different C-terminal and / or N-terminal amino acids).

[0046] Intermediate stringency conditions: The term "intermediate stringency conditions" means that for probes of at least 100 nucleotides in length, pre-hybridize and hybridize at 42°C in 5×SSPE, 0.3% SDS, 200 micrograms / ml shear and denatured salmon sperm DNA, and 35% formamide, followed by a standard Southern blotting procedure for 12-24 hours. Finally, wash the carrier material three times each with 2×SSC and 0.2% SDS at 55°C for 15 minutes.

[0047] Medium to high stringency conditions: The term "medium to high stringency conditions" means that for probes of at least 100 nucleotides in length, pre-hybridize and hybridize at 42°C in 5×SSPE, 0.3% SDS, 200 micrograms / ml shear and denatured salmon sperm DNA, and 35% formamide, followed by a standard Southern blotting procedure for 12-24 hours. Finally, wash the carrier material three times each with 2×SSC and 0.2% SDS at 60°C for 15 minutes.

[0048] Modification: In the context of the polypeptides of the present invention, the term "modification" means that one or more amino acids in the reference amino acid sequence (e.g., SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14) have been altered by substitution with a different amino acid, by insertion of an amino acid, or by deletion, preferably by at least one deletion. The terms "modification," "alteration," and "mutation" are interchangeable and have the same meaning and purpose.

[0049] Mutant: The term "mutant" refers to a polynucleotide that codes for a variant.

[0050] Natural: The term "natural" refers to nucleic acids or polypeptides that are naturally present in host cells.

[0051] Nucleic acid construct: The term “nucleic acid construct” means a single-stranded or double-stranded nucleic acid molecule that is isolated from a naturally occurring gene, modified to contain a nucleic acid segment in a manner not normally found in nature, or a synthetic compound containing one or more regulatory sequences. The term “nucleic acid construct” is synonymous with the term “expression cassette” if the nucleic acid construct contains regulatory sequences required for the expression of the coding sequence of the present invention.

[0052] Operablely linked: The term "operable linked" refers to a configuration in which the control sequence is positioned appropriately relative to the coding sequence of a polynucleotide so that the control sequence induces the expression of the coding sequence.

[0053] Parent or Parent α-Amylase: The term “parent” α-amylase, as used herein, means an α-amylase that has been modified to produce the mutant α-amylase of the present invention. This term also refers to the polypeptide compared to the mutant of the present invention. The parent may be a naturally occurring (wild-type) polypeptide or a mutant thereof prepared by any preferred means. For example, the parent protein may be a mutant of a naturally occurring polypeptide in which the amino acid sequence has been altered or changed. Thus, the parent α-amylase may have one or more (or one or more) amino acid substitutions, deletions, and / or insertions. Thus, the parent α-amylase may be a mutant of the parent α-amylase. The parent may also be an allele variant that is a polypeptide encoded by one of two or more alternative forms of a gene occupying the same chromosomal locus. The terms “parent” or “parent α-amylase” as used herein refer to any α-amylase having at least 60% sequence homology with any of the α-amylases of SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14, or any α-amylase having at least 60% sequence homology with any of the polypeptides of SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14. The parent α-amylase may also be a polypeptide containing the fragments of SEQ ID NOs: 1 to 14. The parent may be a bacterial or fungal α-amylase, preferably a bacterial α-amylase. The amylase having SEQ ID NOs: 1, 2, or 3 may, for example, be the parent or hybrid polypeptide essential to the present invention.

[0054] Purified: The term "purified" means a nucleic acid or polypeptide that is substantially free of other components when measured by analytical techniques well known in the art (for example, a purified polypeptide or nucleic acid may form a separated band in an electrophoretic gel, in a chromatographic eluate, and / or in a culture medium subjected to density gradient centrifugation). A purified nucleic acid or polypeptide is at least about 50% pure, and typically at least about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 99.5%, about 99.6%, about 99.7%, or about 99.8% or more pure (e.g., by weight percentage on a molar basis). In a related sense, a composition is concentrated if the concentration of molecules increases substantially after the application of a purification or concentration technique. The term "concentrated" refers to compounds, polypeptides, cells, nucleic acids, amino acids, or other specific materials or components present in the composition at a higher relative or absolute concentration than that of the starting composition.

[0055] Recombinant: When used in relation to cells, nucleic acids, proteins, or vectors, the term "recombinant" means that they have been modified from their natural state. Therefore, for example, a recombinant cell expresses a gene not found in the cell's natural form (non-recombinant), or expresses a natural gene at a different level or under different conditions than those found in nature. A recombinant nucleic acid differs from its natural sequence by one or more nucleotides and / or is operably ligated to a heterologous sequence, such as a heterologous promoter in an expression vector. A recombinant protein may differ from its natural sequence by one or more amino acids and / or is fused to a heterologous sequence. A vector containing nucleic acids encoding a polypeptide is a recombinant vector. The term "recombinant" is synonymous with "genetically modified" and "gene-transferred."

[0056] Sequence homology: The correlation between two amino acid sequences or two nucleotide sequences is described by a parameter called "sequence homology."

[0057] For the purposes of this invention, sequence homology between two amino acid sequences is determined, preferably using the Needleman-Wunsch algorithm (Needleman and Wunsch, 1970, J.Mol.Biol.48;443-453), as implemented in the Needle program of the EMBOSS package (EMBOSS; The European Molecular Biology Open Software Suite, Rice et al., 2000, Trends Genet.16;276-277) version 5.0.0 or later. The parameters used may be a gap start penalty of 10, a gap extension penalty of 0.5, and an EBLOSUM62 (EMBOSS version of BLOSUM62) permutation matrix. The Needle output labeled "longest identity" (obtained using the -nobrief option) is used as the identity percentage and is calculated as follows: (Number of identical residues × 100) / (Length of alignment - Total number of gaps in alignment)

[0058] For the purposes of this invention, the sequence homology between two deoxyribonucleotide sequences is preferably determined using the Needleman-Wunsch algorithm (Needleman and Wunsch, 1970, cited above), as implemented in the Needle program of the EMBOSS package (EMBOSS; The European Molecular Biology Open Software Suite, Rice et al., 2000, cited above) version 5.0.0 or later. The parameters used are a gap start penalty of 10, a gap extension penalty of 0.5, and the EDNAFULL (EMBOSS version of NCBI NUC4.4) substitution matrix. The Needle output labeled "longest identity" (obtained using the -nobrief option) is used as the identity percentage and is calculated as follows: (100 identical deoxyribonucleotides) / (length of alignment - total number of gaps in the alignment).

[0059] Signal peptide: The term "signal peptide" is defined herein as a peptide in frame (fused) to the amino terminus of a biologically active polypeptide that directs the polypeptide to a cellular secretory pathway. The signal sequence may be determined using techniques known in the art (see, for example, Zhang and Henzel, 2004, Protein Science 13; 2819-2824).

[0060] Starch-binding domain: The terms “starch-binding domain” (SBD) or “carbohydrate-binding module” (CBM) are used interchangeably herein. SBDs can be divided into nine CBM families. Starch is broken down as an energy source by a large number of different amylose-degrading enzymes. However, only about 10 percent of them can bind to and break down raw starch. These enzymes typically have a distinct sequence-structural module called a starch-binding domain that mediates binding to starch granules. An SBD refers to an amino acid sequence that preferentially binds to starch (polysaccharide) substrates or maltosaccharides, α, β, and γ-cyclodextrins, etc. These are typically motifs of about 100 amino acid residues found in about 10 percent of microbial amylose-degrading enzymes.

[0061] Subsequence: The term "subsequence" refers to a polynucleotide having one or more (e.g., several) nucleotides deleted from the 5' and / or 3' ends of the coding sequence of a mature polypeptide, where the subsequence encodes a fragment having α-amylase activity.

[0062] Benefits of Fabric Care: As used herein, the term “benefits of fabric care” is defined as anything not directly related to catalytic stain removal or prevention of dirt re-adhesion, and is also important for the benefits of enzymatic cleaning power. Examples of such benefits of fabric care include preventing or reducing dye transfer from one fabric to another or to a different part of the same fabric (dye transfer suppression), reducing the tendency for pilling by removing protruding or broken fibers from the fabric surface, or removing existing pilling or fuzz (anti-pilling), improving the flexibility of the fabric, clarifying the color of the fabric, and removing particulate matter trapped in the fabric fibers. Enzymatic bleaching is a further benefit of enzymatic cleaning power, and generally, catalytic activity is used to catalyze the formation of bleaching components such as hydrogen peroxide or other peroxides, or bleaching agents.

[0063] Mutants: As used herein, the term "mutant" means a polypeptide having α-amylase activity that includes modifications, i.e., substitutions, insertions, and / or deletions, at one or more (e.g., several) positions. Substitution means replacing an amino acid occupying a position with a different amino acid, deletion means removing an amino acid occupying a position, and insertion means adding an amino acid adjacent to and immediately following an amino acid occupying a position. Mutants have at least 20% of the α-amylase activity of one or more polypeptides of SEQ ID NOs: 1-14, e.g., at least 40%, at least 50%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, but less than 100%.

[0064] Very high stringency conditions: The term "very high stringency conditions" means that for probes of at least 100 nucleotides in length, pre-hybridize and hybridize at 42°C in 5×SSPE, 0.3% SDS, 200 micrograms / ml shear and denatured salmon sperm DNA, and 50% formamide, followed by a standard Southern blotting procedure for 12-24 hours. Finally, wash the carrier material three times each with 2×SSC and 0.2% SDS at 70°C for 15 minutes.

[0065] Very low stringency conditions: The term "very low stringency conditions" means that for probes of at least 100 nucleotides in length, pre-hybridize and hybridize in 5×SSPE, 0.3% SDS, 200 micrograms / ml shear and denatured salmon sperm DNA, and 25% formamide at 42°C, followed by a standard Southern blotting procedure for 12-24 hours. Finally, wash the carrier material three times each with 2×SSC and 0.2% SDS at 45°C for 15 minutes.

[0066] Cleaning Performance: In the context of this specification, the term “cleaning performance” is used to describe the ability of an enzyme to remove starch or starch-containing stains present on an object to be cleaned, for example, when cleaning hard surfaces such as laundry or dishwashing. The term “cleaning performance” generally includes cleaning hard surfaces such as dishwashing, but also includes cleaning performance on textiles such as laundry, and cleaning performance in industrial and commercial cleaning. Cleaning performance may be quantified by calculating a so-called intensity value.

[0067] Wild-type enzyme: The term "wild-type" α-amylase refers to α-amylase expressed by naturally occurring microorganisms such as bacteria, yeasts, or filamentous fungi found in nature. The terms "wild-type enzyme" and "parent enzyme" can be used interchangeably when the parent enzyme is not a mutant enzyme.

[0068] nomenclature For the purposes of this invention, the nomenclature [Y / F] means that the amino acid at that position may be tyrosine (Try, Y) or phenylalanine (Phe, F). Similarly, in other combinations as described herein, the nomenclature [V / G / A / I] means that the amino acid at that position may be valine (Val, V), glycine (Gly, G), alanine (Ala, A), or isoleucine (Ile, I), etc. Amino acid X is defined to be any of the 20 natural amino acids unless otherwise specified.

[0069] Conventional methods for notating variants For the purposes of the present invention, the polypeptide disclosed in SEQ ID NO: 1 is used to determine the corresponding amino acid residue in another α-amylase. Alternatively, the polypeptide disclosed in SEQ ID NO: 2 or SEQ ID NO: 3 may be used to determine the corresponding amino acid residue in another α-amylase. The amino acid sequence of the other α-amylase is aligned with the polypeptide disclosed in SEQ ID NO: 1, and based on this alignment, the amino acid position number corresponding to any amino acid residue in the polypeptide disclosed in SEQ ID NO: 2 or SEQ ID NO: 3 is determined, preferably using the Needleman-Wunsch algorithm (Needleman and Wunsch, 1970, J.Mol.Biol.48;443-453), as implemented in the Needle program of the EMBOSS package (EMBOSS; The European Molecular Biology Open Software Suite, Rice et al., 2000, Trends Genet.16;276-277) version 3.0.0 or later.

[0070] Identification of corresponding amino acid residues within other α-amylases is done using MUSCLE (multiple sequence comparison by log-expectation; version 3.5 and later; Edgar, 2004, Nucleic Acids Research 32; 1792-1797), MAFFT (version 6.857 and later; Katoh and Kuma, 2002, Nucleic Acids Research 30; 3059-3066; Katoh et al., 2005, Nucleic Acids Research 33; 511-518; Katoh and Toh, 2007, Bioinformatics 23; 372-374; Katoh et al., 2009, Methods in Molecular Biology 537; 39-64; Katoh and Toh, 2010, Bioinformatics 26; 1899-1900), and EMBOSS. One example is ClustalW (version 1.83 and later; Thompson et al., 1994, Nucleic Acids Research 22; 4673-4680), which uses EMMA, but it can also be determined by aligning multiple polypeptide sequences using several computer programs with their corresponding default parameters, not limited to these.

[0071] If other enzymes branch off from the polypeptide of SEQ ID NO: 1, and as a result their relationship is not detected by conventional sequence-based comparisons (Lindahl and Elofsson, 2000, J.Mol.Biol.295;613-615), other pairwise sequence comparison algorithms can be used. Using search programs that utilize probabilistic representations (profiles) of polypeptide families to search databases can achieve higher sensitivity in sequence-based searches. For example, the PSI-BLAST program can generate profiles through an iterative database search process to detect distant homologs (Atschul et al., 1997, Nucleic Acids Res.25;3389-3402).

[0072] Higher sensitivity can be achieved if a polypeptide family or superfamily has one or more samples in the protein structure database. Programs such as GenTHREADER (Jones, 1999, J.Mol.Biol.287;797-815; McGuffin and Jones, 2003, Bioinformatics 19;874-881) utilize information from various sources (PSI-BLAST, secondary structure prediction, structural alignment profiles, solvation potential) as input to a neural network that predicts the structural folding of the query sequence. Similarly, the method of Gough et al., 2000, J.Mol.Biol.313;903-919 can be used to align the sequence of an unknown structure with a superfamily model present in the SCOP database. These alignments can then be used to generate polypeptide homology models, and the accuracy of such models can be evaluated using various tools developed for that purpose.

[0073] For proteins with known structures, several tools and resources are available for searching and generating structural alignments. For example, the SCOP superfamily of proteins is structurally aligned, and these alignments are available and downloadable. Two or more protein structures can be aligned using various algorithms such as distance alignment matrices (Holm and Sander, 1998, Proteins 33;88-96) or combinatorial extensions (hindyalov and Bourne, 1998, Protein Engineering 11;739-747), and implementations of these algorithms can be used to query structural databases containing the target structure to find possible structural congeners (e.g., Holm and Park, 2000, Bioinformatics 16;566-567).

[0074] When describing variants, the following nomenclature will be applied for ease of reference. Accepted IUPAC one- or three-letter amino acid abbreviations will be used.

[0075] Substitutions: For amino acid substitutions, the following nomenclature is used: original amino acid, position, and substituted amino acid. Therefore, the substitution of threonine at position 226 with alanine is written as "Thr226Ala" or "T226A". If the amino acid at a given position can be substituted with any other amino acid, it is written as T226ACDEFGHIKLMNPQRSWVY. This means that the threonine at position 226 can be substituted with an amino acid selected from the group consisting of A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, W, V, or Y. Similarly, if the amino acid at a given position can be substituted with one amino acid selected from a specific group of amino acids, for example, if the threonine at position 226 can be substituted with tyrosine, phenylalanine, or histidine, it is written as T226YFH. Furthermore, different modifications at a given position may be separated by commas. For example, "Arg170Tyr,Glu" or "R170Y,E" indicate that the arginine at position 170 has been replaced with tyrosine or glutamic acid. Thus, "Tyr167Gly,Ala+Arg170Gly,Ala" represents the following variants: "Tyr167Gly+Arg170Gly", "Tyr167Gly+Arg170Ala", "Tyr167Ala+Arg170Gly", and "Tyr167Ala+Arg170Ala".

[0076] Multiple mutations are distinguished by an addition sign ("+"). For example, "Gly205Arg+Ser411Phe" or "G205R+S411F" indicate the substitution of glycine (G) at position 205 with arginine (R) and serine (S) at position 411 with phenylalanine (F).

[0077] Deletion. In the case of amino acid deletions, the following nomenclature is used: original amino acid, position, and *. Therefore, a deletion of glycine at position 195 is written as "Gly195*" or "G195*". Multiple deletions are separated by an addition sign ("+"), for example, "Gly195*+Ser411*" or "G195*+S411*".

[0078] Insertion. In the case of amino acid insertion, the following nomenclature is used: original amino acid, position, original amino acid, and inserted amino acid. Therefore, the insertion of lysine after glycine at position 195 is written as "Gly195GlyLys" or "G195GK". Insertions of multiple amino acids are written as [original amino acid, position, original amino acid, first inserted amino acid, second inserted amino acid...]. For example, the insertion of lysine and alanine after glycine at position 195 is written as "Gly195GlyLysAla" or "G195GKA".

[0079] In such cases, the inserted amino acid residues are numbered by adding a lowercase letter to the position number of the amino acid residue preceding the inserted amino acid residue. Therefore, in the example above, the sequence would be as follows:

[0080] [Table 1]

[0081] Multiple modifications. Mutants containing multiple modifications are separated by the addition sign "+". For example, "Arg170Tyr+Gly195Glu" or "R170Y+G195E" indicates that arginine and glycine at positions 170 and 195 are replaced with tyrosine and glutamic acid, respectively. [Modes for carrying out the invention]

[0082] Parent α-amylase The present invention relates to a polypeptide having amylase activity, wherein the polypeptide is a hybrid polypeptide.

[0083] The present invention provides a polypeptide having α-amylase activity, comprising an A domain having at least 60% sequence homology with the A domain of any of SEQ ID NOs: 1, 2, 3, 4, or 5; a B domain having at least 60% sequence homology with the B domain of any of SEQ ID NOs: 1, 2, 10, 11, or 12; and a C domain having at least 60% sequence homology with the C domain of any of SEQ ID NOs: 1, 2, 3, 4, or 5.

[0084] The present invention includes a hybrid α-amylase of a parent polypeptide having α-amylase activity, comprising an A domain, a B domain, and a C domain, wherein the hybrid is a fusion polypeptide.

[0085] In one embodiment, the A domain has at least 60% sequence homology with the A domain of Sequence ID No. 1, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology.

[0086] In one embodiment, the A domain has at least 60% sequence homology with the A domain of SEQ ID NO: 2, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology.

[0087] In one embodiment, the A domain has at least 60% sequence homology with the A domain of Sequence ID No. 3, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology.

[0088] In one embodiment, the A domain has at least 60% sequence homology with the A domain of sequence number 4, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology.

[0089] In one embodiment, the A domain has at least 60% sequence homology with the A domain of sequence number 5, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology.

[0090] In one embodiment, the B domain has at least 60% sequence homology with the B domain of Sequence ID No. 1, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology.

[0091] In one embodiment, the B domain has at least 60% sequence homology with the B domain of Sequence ID No. 2, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology.

[0092] In one embodiment, the B domain has at least 60% sequence homology with the B domain of sequence number 10, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology.

[0093] In one embodiment, the B domain has at least 60% sequence homology with the B domain of Sequence ID No. 11, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology.

[0094] In one embodiment, the B domain has at least 60% sequence homology with the B domain of sequence number 12, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology.

[0095] In one embodiment, the C domain has at least 60% sequence homology with the C domain of SEQ ID NO: 1, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology.

[0096] In one embodiment, the C domain has at least 60% sequence homology with the C domain of SEQ ID NO: 2, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology.

[0097] In one embodiment, the C domain has at least 60% sequence homology with the C domain of SEQ ID NO: 3, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology.

[0098] In one embodiment, the C domain has at least 60% sequence homology with the C domain of SEQ ID NO: 4, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology.

[0099] In one embodiment, the C domain has at least 60% sequence homology with the C domain of SEQ ID NO: 5, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology.

[0100] α-amylase may be produced by substituting the B domain or a portion thereof of one α-amylase with the B domain or a portion thereof of another α-amylase. α-amylase may also be produced by substituting the A domain and C domain or a portion thereof of one α-amylase with the A domain and C domain or a portion thereof of another α-amylase.

[0101] In the case of Bacillus α-amylase (SEQ ID NO: 4), whose B domain is determined to be amino acid residues 109-206, the sequence to be substituted with the corresponding sequence of another α-amylase is one that starts at positions 93-113 and ends at positions 195-215 of SEQ ID NO: 12, for example, starting at positions 97-109 and ending at positions 199-215, or starting at positions 100-106 and ending at positions 202-208, particularly positions 109-206. The A and C domains of Bacillus α-amylase were determined to be amino acid residues 1-108 (A1) + 207-396 (A2) and 397-485, respectively. The α-amylase of the present invention may include an A1 domain that starts at positions 1-5 of SEQ ID NO: 4 and ends at positions 94-114, for example, starting at positions 1-3 and ending at positions 99-110, or starting at positions 1-3 and ending at positions 104-109, particularly positions 1-108. The α-amylase of the present invention may include an A2 domain and a C domain that starts at positions 201-221 of SEQ ID NO: 4 and ends at positions 478-514, for example, starting at positions 203-211 and ending at positions 480-510, or starting at positions 205-216 and ending at positions 482-484, particularly positions 207-485.

[0102] In one embodiment, the hybrid polypeptide comprises or consists of the amino acid sequence described in SEQ ID NO: 1 or 2. In another embodiment, the hybrid polypeptide is an allele variant of the polypeptide of SEQ ID NO: 1 or 2.

[0103] In another embodiment, the parent polypeptide may contain or consist of the amino acid sequence described in SEQ ID NO: 1. As described elsewhere in this specification, the parent polypeptide may be any polypeptide having α-amylase activity and having at least 60% sequence homology to any one of the amino acid sequences described in SEQ ID NOs: 1 to 14.

[0104] In one embodiment, the parent polypeptide has at least 60%, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence homology with the amino acid sequence described in SEQ ID NOs: 2, and this parent polypeptide has α-amylase activity. In one embodiment, the amino acid sequence of the parent polypeptide differs from the amino acid sequences described in SEQ ID NOs: 1 to 14 by up to 10 amino acids, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids.

[0105] According to this invention, a value of 1.0 corresponds to the performance observed in the parent polypeptide. A value greater than 1.0 indicates an improvement in the performance of the tested hybrid compared to the parent polypeptide. Therefore, any value >1.0 indicates an improvement in properties, such as an increase in the specific activity of the hybrid polypeptide compared to the parent polypeptide.

[0106] In one embodiment, the hybrid polypeptide is the parent polypeptide.

[0107] The parent α-amylase may be a polypeptide having sequence homology of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0108] In one embodiment, the parent has sequence homology with the polypeptide of SEQ ID NO: 1 having α-amylase activity of at least 60%, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or 100%. In one embodiment, the amino acid sequence of the parent differs from the polypeptide of SEQ ID NO: 1 by 10 or fewer amino acids, for example, 5 amino acids, 4 amino acids, 3 amino acids, 2 amino acids, and 1 amino acid.

[0109] The parent preferably contains or consists of the amino acid sequence of SEQ ID NO: 1. In one embodiment, the parent contains or consists of the polypeptide of SEQ ID NO: 1. In another embodiment, the parent is an allele variant of the polypeptide of SEQ ID NO: 1.

[0110] In one embodiment, the parent has sequence homology with the polypeptide of SEQ ID NO: 2 having α-amylase activity of at least 60%, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or 100%. In one embodiment, the amino acid sequence of the parent differs from the polypeptide of SEQ ID NO: 2 by 10 or fewer amino acids, for example, 5 amino acids, 4 amino acids, 3 amino acids, 2 amino acids, and 1 amino acid.

[0111] The parent preferably contains or consists of the amino acid sequence of SEQ ID NO: 2. In one embodiment, the parent contains or consists of the polypeptide of SEQ ID NO: 2. In another embodiment, the parent is an allele variant of the polypeptide of SEQ ID NO: 2.

[0112] In one embodiment, the parent has sequence homology with the polypeptide of SEQ ID NO: 3 having α-amylase activity of at least 60%, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or 100%. In one embodiment, the amino acid sequence of the parent differs from the polypeptide of SEQ ID NO: 3 by 10 or fewer amino acids, for example, 5 amino acids, 4 amino acids, 3 amino acids, 2 amino acids, and 1 amino acid.

[0113] The parent preferably contains or consists of the amino acid sequence of SEQ ID NO: 3. In one embodiment, the parent contains or consists of the polypeptide of SEQ ID NO: 3. In another embodiment, the parent is an allele variant of the polypeptide of SEQ ID NO: 3.

[0114] In one embodiment, the parent has sequence homology with the polypeptide of SEQ ID NO: 4 having α-amylase activity of at least 60%, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or 100%. In one embodiment, the amino acid sequence of the parent differs from the polypeptide of SEQ ID NO: 4 by 10 or fewer amino acids, for example, 5 amino acids, 4 amino acids, 3 amino acids, 2 amino acids, and 1 amino acid.

[0115] The parent preferably contains or consists of the amino acid sequence of SEQ ID NO: 4. In one embodiment, the parent contains or consists of the polypeptide of SEQ ID NO: 4. In another embodiment, the parent is an allele variant of the polypeptide of SEQ ID NO: 4.

[0116] In one embodiment, the parent has sequence homology with the polypeptide of SEQ ID NO: 5 having α-amylase activity of at least 60%, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or 100%. In one embodiment, the amino acid sequence of the parent differs from the polypeptide of SEQ ID NO: 5 by 10 or fewer amino acids, for example, 5 amino acids, 4 amino acids, 3 amino acids, 2 amino acids, and 1 amino acid.

[0117] The parent preferably contains or consists of the amino acid sequence of SEQ ID NO: 5. In one embodiment, the parent contains or consists of the polypeptide of SEQ ID NO: 5. In another embodiment, the parent is an allele variant of the polypeptide of SEQ ID NO: 5.

[0118] In one embodiment, the parent has sequence homology with the polypeptide of SEQ ID NO: 6 having α-amylase activity of at least 60%, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or 100%. In one embodiment, the amino acid sequence of the parent differs from the polypeptide of SEQ ID NO: 6 by 10 or fewer amino acids, for example, 5 amino acids, 4 amino acids, 3 amino acids, 2 amino acids, and 1 amino acid.

[0119] The parent preferably contains or consists of the amino acid sequence of SEQ ID NO: 6. In one embodiment, the parent contains or consists of the polypeptide of SEQ ID NO: 6. In another embodiment, the parent is an allele variant of the polypeptide of SEQ ID NO: 6.

[0120] In one embodiment, the parent has sequence homology with the polypeptide of Sequence ID No. 7 having α-amylase activity of at least 60%, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or 100%. In one embodiment, the amino acid sequence of the parent differs from the polypeptide of Sequence ID No. 7 by 10 or fewer amino acids, for example, 5 amino acids, 4 amino acids, 3 amino acids, 2 amino acids, and 1 amino acid.

[0121] The parent preferably contains or consists of the amino acid sequence of SEQ ID NO: 7. In one embodiment, the parent contains or consists of the polypeptide of SEQ ID NO: 7. In another embodiment, the parent is an allele variant of the polypeptide of SEQ ID NO: 7.

[0122] In one embodiment, the parent has sequence homology with the polypeptide of SEQ ID NO: 8 having α-amylase activity of at least 60%, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or 100%. In one embodiment, the amino acid sequence of the parent differs from the polypeptide of SEQ ID NO: 8 by 10 or fewer amino acids, for example, 5 amino acids, 4 amino acids, 3 amino acids, 2 amino acids, and 1 amino acid.

[0123] The parent preferably contains or consists of the amino acid sequence of SEQ ID NO: 8. In one embodiment, the parent contains or consists of the polypeptide of SEQ ID NO: 8. In another embodiment, the parent is an allele variant of the polypeptide of SEQ ID NO: 8.

[0124] In one embodiment, the parent has sequence homology with the polypeptide of Sequence ID No. 9 having α-amylase activity of at least 60%, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or 100%. In one embodiment, the amino acid sequence of the parent differs from the polypeptide of Sequence ID No. 9 by 10 or fewer amino acids, for example, 5 amino acids, 4 amino acids, 3 amino acids, 2 amino acids, and 1 amino acid.

[0125] The parent preferably contains or consists of the amino acid sequence of SEQ ID NO: 9. In one embodiment, the parent contains or consists of the polypeptide of SEQ ID NO: 9. In another embodiment, the parent is an allele variant of the polypeptide of SEQ ID NO: 9.

[0126] In one embodiment, the parent has sequence homology with the polypeptide of SEQ ID NO: 10 having α-amylase activity of at least 60%, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or 100%. In one embodiment, the amino acid sequence of the parent differs from the polypeptide of SEQ ID NO: 10 by 10 or fewer amino acids, for example, 5 amino acids, 4 amino acids, 3 amino acids, 2 amino acids, and 1 amino acid.

[0127] The parent preferably contains or consists of the amino acid sequence of SEQ ID NO: 10. In one embodiment, the parent contains or consists of the polypeptide of SEQ ID NO: 10. In another embodiment, the parent is an allele variant of the polypeptide of SEQ ID NO: 10.

[0128] In one embodiment, the parent has sequence homology with the polypeptide of SEQ ID NO: 11 having α-amylase activity of at least 60%, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or 100%. In one embodiment, the amino acid sequence of the parent differs from the polypeptide of SEQ ID NO: 11 by 10 or fewer amino acids, for example, 5 amino acids, 4 amino acids, 3 amino acids, 2 amino acids, and 1 amino acid.

[0129] The parent preferably contains or consists of the amino acid sequence of SEQ ID NO: 11. In one embodiment, the parent contains or consists of the polypeptide of SEQ ID NO: 11. In another embodiment, the parent is an allele variant of the polypeptide of SEQ ID NO: 11.

[0130] In one embodiment, the parent has sequence homology with the polypeptide of SEQ ID NO: 12 having α-amylase activity of at least 60%, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or 100%. In one embodiment, the amino acid sequence of the parent differs from the polypeptide of SEQ ID NO: 12 by 10 or fewer amino acids, for example, 5 amino acids, 4 amino acids, 3 amino acids, 2 amino acids, and 1 amino acid.

[0131] The parent preferably contains or consists of the amino acid sequence of SEQ ID NO: 12. In one embodiment, the parent contains or consists of the polypeptide of SEQ ID NO: 12. In another embodiment, the parent is an allele variant of the polypeptide of SEQ ID NO: 12.

[0132] In one embodiment, the parent has sequence homology with the polypeptide of SEQ ID NO: 13 having α-amylase activity of at least 60%, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or 100%. In one embodiment, the amino acid sequence of the parent differs from the polypeptide of SEQ ID NO: 13 by 10 or fewer amino acids, for example, 5 amino acids, 4 amino acids, 3 amino acids, 2 amino acids, and 1 amino acid.

[0133] The parent preferably contains or consists of the amino acid sequence of SEQ ID NO: 13. In one embodiment, the parent contains or consists of the polypeptide of SEQ ID NO: 13. In another embodiment, the parent is an allele variant of the polypeptide of SEQ ID NO: 13.

[0134] In one embodiment, the parent has sequence homology with the polypeptide of SEQ ID NO: 14 having α-amylase activity of at least 60%, for example, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, or 100%. In one embodiment, the amino acid sequence of the parent differs from the polypeptide of SEQ ID NO: 14 by 10 or fewer amino acids, for example, 5 amino acids, 4 amino acids, 3 amino acids, 2 amino acids, and 1 amino acid.

[0135] The parent preferably contains or consists of the amino acid sequence of SEQ ID NO: 14. In one embodiment, the parent contains or consists of the polypeptide of SEQ ID NO: 14. In another embodiment, the parent is an allele variant of the polypeptide of SEQ ID NO: 14.

[0136] Nucleic acid probes may be designed using the amino acid sequences of SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, and 14, or fragments thereof, and the DNA encoding the parent may be identified and cloned from strains of different genera or species according to methods well known in the art. In particular, such probes may be hybridized with the genome or cDNA of the target genus or species according to a standard Southern blotting procedure to identify and isolate the corresponding genes. Such probes may be significantly shorter than the entire sequence, but must be at least 14, for example, at least 25, at least 35, or at least 70 nucleotides long. Preferably, the nucleic acid probe is at least 100 nucleotides long, for example, at least 200, at least 300, at least 400, at least 500, at least 600, at least 700, at least 800, or at least 900 nucleotides long. Both DNA probes and RNA probes can be used. Probes are typically labeled to detect the corresponding gene (e.g., 32 P, 3 H, 35 (S, biotin, or avidin). Such probes are included in the present invention.

[0137] Genomic DNA or cDNA libraries prepared from such other strains may be hybridized with the probes described above and screened for whether they encode the parent. The genome or other DNA of such other strains may be isolated by agarose or polyacrylamide gel electrophoresis or other separation techniques. The DNA from the library or the isolated DNA may be transferred and immobilized on nitrocellulose or other suitable carrier material. This carrier material is used in Southern blotting to identify clones or DNA that hybridize with the parent or a portion thereof.

[0138] For the purposes of the present invention, hybridization indicates that a polynucleotide hybridizes under low to very high stringency conditions to a labeled nucleotide probe corresponding to a polynucleotide encoding SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or a subsequence thereof. The molecule hybridized by the probe can be detected, for example, using an X-ray film or any other detection means known in the art.

[0139] In one embodiment, the nucleic acid probe is a polynucleotide encoding the polypeptide of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or a fragment thereof.

[0140] For long probes of at least 100 nucleotides in length, very low to very high stringency conditions are defined as pre-hybridizing and hybridizing at 42°C in 5×SSPE, 0.3% SDS, 200 micrograms / ml shear and denatured salmon sperm DNA, 25% formamide for very low and low stringency, 35% formamide for medium and medium-high stringency, or 50% formamide for high and very high stringency, followed by a standard Southern blotting procedure for optimally 12-24 hours. Finally, the carrier material is washed three times each for 15 minutes at 45°C (very low stringency), 50°C (low stringency), 55°C (medium stringency), 60°C (medium-high stringency), 65°C (high stringency), or 70°C (very high stringency) using 2×SSC, 0.2% SDS.

[0141] For short probes approximately 15 to 70 nucleotides in length, stringency conditions are defined as pre-hybridizing and hybridizing at approximately 5°C to 10°C below the Tm calculated using calculations by Bolton and McCarthy (1962, Proc. Natl. Acad. Sci. USA 48; 1390) in 0.9 M NaCl, 0.09 M Tris-HCl (pH 7.6), 6 mM EDTA, 0.5% NP-40, 1 × Denhart solution, 1 mM sodium pyrophosphate, 1 mM sodium dihydrogen phosphate, 0.1 mM ATP, and 0.2 mg of yeast RNA per ml, followed by standard Southern blotting procedures, optimally for 12 to 24 hours. Finally, the carrier material was washed once for 15 minutes in 0.1% SDS in addition to 6×SCC, and then twice for 15 minutes each using 6×SSC at 5°C to 10°C below the calculated Tm.

[0142] The parent may be obtained from any genus of microorganism. For the purposes of this invention, the term “obtained from,” when used herein in relation to a given source, means that the parent encoded by polynucleotides is produced by the source or by a strain into which polynucleotides from the source have been inserted. In one embodiment, the parent is secreted extracellularly.

[0143] The parent can be a bacterial α-amylase. For example, the parent may be a Gram-positive α-amylase from the genera Bacillus, Clostridium, Enterococcus, Geobacillus, Lactobacillus, Lactococcus, Oceanobacillus, Staphylococcus, Streptococcus, or Streptomyces. The material may be a bacterial polypeptide, or a Gram-negative bacterial polypeptide such as α-amylase from Campylobacter, Escherichia coli, Flavobacterium, Fusobacterium, Helicobacter, Ilyobacter, Neisseria, Pseudomonas, Salmonella, or Ureaplasma.

[0144] In one manifestation, the parents are Bacillus alkalophilus, Bacillus amyloliquefaciens, Bacillus brevis, Bacillus circulans, Bacillus clausii, Bacillus coagulans, Bacillus firmus, Bacillus lautus, Bacillus lentus, Bacillus licheniformis, Bacillus megaterium, and Bacillus pmyrus. It is the α-amylase of Bacillus pumilus, Bacillus stearothermophilus, Bacillus subtilis, or Bacillus thuringiensis.

[0145] In another embodiment, the parent is the α-amylase of Streptococcus equisimilis, Streptococcus pyogenes, Streptococcus uberis, or Streptococcus equi subsp. Zooepidemicus.

[0146] In another embodiment, the parent is the α-amylase of Streptomyces achromogenes, Streptomyces avermitilis, Streptomyces coelicolor, Streptomyces griseus, or Streptomyces lividans.

[0147] In another embodiment, the parent is an α-amylase of the genus Bacillus, for example, the α-amylase of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, or SEQ ID NO: 14.

[0148] With respect to the species mentioned above, it will be understood that the present invention encompasses both the complete and imperfect generations, regardless of the species' name, as well as other taxonomic equivalents, such as anamorphs. Those skilled in the art will readily understand the identity of appropriate equivalents.

[0149] Strains of these species are publicly and readily available at numerous microbial strain preservation institutions, including the American Type Culture Collection (ATCC), Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH (DSM), Centraalbureau Voor Schimmelcultures (CBS), and the Agricultural Research Service Patent Culture Collection, Northern Regional Research Center (NRRL).

[0150] The parent may be identified and obtained from other sources, including microorganisms isolated from nature (e.g., soil, compost, water, etc.) or DNA samples obtained directly from natural substances (e.g., soil, compost, water, etc.), using the probe described above. Methods for directly isolating microorganisms and DNA from natural habitats are well known in the art. The polynucleotide encoding the parent may then be extracted by similarly screening a genomic library or cDNA library of another microorganism or mixed DNA sample. Once the polynucleotide encoding the parent has been detected by the probe, it may be isolated and cleaned using methods known to those skilled in the art (see, for example, Sambrook et al., 1989 (cited above)).

[0151] The parent may be a hybrid polypeptide in which a portion of one polypeptide is fused to the N-terminus or C-terminus of a portion of another polypeptide.

[0152] The parent polypeptide may also be a fusion polypeptide or a cleavable fusion polypeptide in which one polypeptide is fused to another polypeptide at the N-terminus or C-terminus. The fusion polypeptide is produced by fusing a polynucleotide encoding one polypeptide to a polynucleotide encoding another polypeptide. Methods for producing fusion polypeptides are known in the art and include ligating the coding sequences encoding polypeptides such that they are in-frame and the expression of the fusion polypeptide is under the control of the same promoter and terminator. The fusion protein may also be constructed using intein techniques in which the fusion occurs post-translation (Cooper et al., 1993, EMBO J.12; 2575-2583; Dawson et al., 1994, Science 266; 776-779).

[0153] The fusion polypeptide can also contain a cleavage site between the two polypeptides. When the fusion protein is secreted, this site is cleaved, releasing the two polypeptides. Examples of cleavage sites include Martin et al.,2003,J.Ind.Microbiol.Biotechnol.3;568-576;Svetina et al.,2000,J.Biotechnol.76;245-251;Rasmussen-Wilson et al. al.,1997,Appl.Environ.Microbiol.63;3488-3493;Ward et al.,1995,Biotechnology 13;498-503;and Contreras et al.,1991,Biotechnology 9;378-381;Eaton et al.,1986,Biochemistry 25;505-512;Collins-Racie et al.,1995,Biotechnology 13;982-987;Carter et al. Examples of sites disclosed include, but are not limited to, those described in al., 1989, Proteins: Structure, Function, and Genetics 6;240-248; and Stevens, 2003, Drug Discovery World 4;35-48.

[0154] Preparation of mutants The present invention (a) uses sequence number 1 for numbering purposes to 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 1 37, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 25 6, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 361, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 47 The present invention relates to a method for obtaining a mutant having α-amylase activity, comprising: (a) introducing a modification at one or more positions corresponding to positions 0, 472, 473, 474, 475, and 476 into a parent α-amylase, wherein each modification is independently a substitution or deletion, and the mutant has α-amylase activity; and (b) recovering the mutant.

[0155] A method for obtaining a mutant having α-amylase activity is (a) an amino acid sequence as described in SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126 ,128,131,132,134,135,136,137,138,140,142,144,146,151,152,155,159,160,165,167,169,172,173,174,176,177,178,180,181,182,183,185,188,189,193,194,196,202,203,205 ,224,225,242,243,244,252,256,258,259,260,261,266,276,279,280,283,286,302,303,315,320,321,322,323,346,354,359,360,361,362,365,371,383,390,394,395,405,409,41 (b) introducing a modification to a parent α-amylase at one or more positions corresponding to positions 4, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein each modification is independently a substitution or deletion, and the mutant has α-amylase activity; and (b) recovering the mutant.

[0156] In one embodiment, a method for obtaining a mutant having α-amylase activity is (a) using SEQ ID NO: 1 for numbering, the amino acid sequences 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106 as described in SEQ ID NOs: 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 or 14 ,109,110,111,113,114,116,117,118,123,126,128,131,132,134,135,136,137,138,140,142,144,146,151,152,155,159,160,165,167,169,172,173,174,176,177,178,180,181,182,183,185,188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 361, 362, 365, 371, 383, 390, 394, 3 (b) introducing a modification to a parent α-amylase at one or more positions corresponding to positions 95, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein each modification is independently a substitution or deletion, and the mutant has α-amylase activity; and (b) recovering the mutant.

[0157] In one embodiment, the modification is a substitution. In another embodiment, the modification is a deletion.

[0158] In another embodiment, a method for obtaining a mutant having α-amylase activity is (a) introducing a modification at one or more positions into the parent α-amylase, wherein the modification is H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N1 06D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F116L, F116 N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T131I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W1 40Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F155W, W159H , W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S, E169I, E 169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K178A, R18 0*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T182G, T182I , T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N, A185Q, A1 85R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L205N, L205 Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243Q, F244I, F 244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F261K, F26 1P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N280Y, W283N,S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y, M322G, M322Y, H323A, H323N, H323W, W346N, L 354V, E359W, Q360M, Q360W, Q360Y, G361M, Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F3 (b) introducing a mutant selected from 95Y, D405G, I409L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E, and G476A, and (b) recovering the mutant.

[0159] In another embodiment, a method for obtaining a mutant having α-amylase activity is (a) introducing a modification at one or more positions into the parent α-amylase, wherein the modification is N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L of the amino acid sequence described in SEQ ID NOs. 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 or 14, using SEQ ID NO 1 for numbering. , V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A 111L, G113Q, G113S, G113A, T114K, F116L, F116N, F116Q, F116W, V117G, D118 Q, D118T, D123N, N126D, N128Y, T131I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D14 4N, P146S, P146H, T151A, Y152L, F155W, W159H, W159L, Y160P, T165V, W167A , W167D, W167E, W167F, W167P, W167S, E169I, E169Q, E169V, K172G, K172Q, L1 73F, N174K, N174S, I176V, Y177D, K178A, R180Q, R180S, S181L, S181T, T182 D, T182E, T182G, T182I, T182L, T182M, T182S, G183P, A185D, A185E, A185K, A 185N, A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L20 5F, L205N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M26 0W, F261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W,N280Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y, M322G, M322Y, H323A, H323N, H323 W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361M, Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N3 (b) introducing a mutant selected from 94D, F395Y, D405G, I409L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E, and G476A, and (b) recovering the mutant.

[0160] In one embodiment, the method comprises introducing a deletion at one or more positions into a parent α-amylase, the deletion being selected from H1*, H2*, R180*, S181*, T182* and G183* of polypeptides of sequence numbers 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 or 14, using sequence number 1 for numbering purposes, and recovering the mutant.

[0161] Mutants may be prepared using any mutagenesis method known in the art, such as site-directed mutagenesis, synthetic gene construction methods, semi-synthetic gene construction methods, random mutagenesis, shuffling procedures, etc.

[0162] Site-directed mutagenesis is a technique that induces one or more mutations at one or more defined sites within a polynucleotide that codes for a parent.

[0163] Site-directed mutagenesis can be performed in vitro by PCR, which involves using oligonucleotide primers containing the desired mutation. Site-directed mutagenesis can also be performed in vitro by cassette mutagenesis, which involves cleaving a site in a plasmid containing a parent-coding polynucleotide with a restriction enzyme, and then ligating the oligonucleotide containing the mutation into the polynucleotide. Typically, the restriction enzyme digested by the plasmid and the oligonucleotide are the same, which allows the adherent ends of the plasmid and insert to bind to each other. See, for example, Scherer and Davis, 1979, Proc. Natl. Acad. Sci. USA 76;4949-4955; and Barton et al., 1990, Nucleic Acids Res. 18;7349-4966.

[0164] Site-directed mutagenesis can also be performed in vivo using methods known in the art. See, for example, U.S. Patent Application Publication No. 2004 / 0171154, Storici et al., 2001, Nature Biotechnol. 19; 773-776; Kren et al., 1998, Nat. Med. 4; 285-290; and Calissano and Macino, 1996, Fungal Genet. Newslett. 43; 15-16.

[0165] Any site-directed mutagenesis procedure may be used in the present invention. Many commercially available kits exist that can be used to prepare mutants.

[0166] Synthetic gene construction methods require the in vitro synthesis of polynucleotide molecules designed to encode the target polypeptide. Gene synthesis can be performed using a variety of techniques, such as the multi-microchip-based technology described by Tian et al. (2004, Nature 432:1050-1054), and similar techniques in which oligonucleotides are synthesized and assembled on photoprogrammable microfluidic chips.

[0167] Single or multiple amino acid substitutions, deletions, and / or insertions can be generated and tested using known methods of mutagenesis, recombination, and / or shuffling, and then relevant screening procedures can be performed as disclosed in Reidhaar-Olson and Sauer, 1988, Science 241;53-57; Bowie and Sauer, 1989, Proc. Natl. Acad. Sci. USA 86;2152-2156; International Publication No. 95 / 17413; or International Publication No. 95 / 22625. Other available methods include error-prone PCR, phage display (e.g., Lowman et al., 1991, Biochemistry 30;10832-10837; U.S. Patent No. 5,223,409; International Publication No. 92 / 06204), and region-specific mutagenesis (Derbyshire et al., 1986, Gene 46;145; Ner et al., 1988, DNA 7;127).

[0168] Mutagenesis / shuffling methods, when combined with high-throughput automated screening methods, can detect the activity of cloned, mutagenesized polypeptides expressed by host cells (Ness et al., 1999, Nature Biotechnology 17;893-896). Mutagenesized DNA molecules encoding active polypeptides can be recovered from host cells and readily sequenced using standard methods in the art. These methods allow for rapid determination of the importance of individual amino acid residues within the polypeptide.

[0169] Semi-synthetic gene construction methods are carried out by combining synthetic gene construction methods and / or site-directed mutagenesis and / or random mutagenesis and / or shuffling methods. Typical semi-synthetic construction methods include processes using synthesized polynucleotide fragments in combination with PCR techniques. Thus, a defined region of the gene may be newly synthesized, while other regions may be amplified using site-directed mutagenesis primers, and other regions may be subjected to error-prone PCR or non-error-prone PCR amplification. Subsequently, the polynucleotide subsequence may be shuffled.

[0170] α-amylase mutant The present invention uses sequence number 1 for numbering purposes to represent 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 13 2, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 361, 362, 365, 371, 38 This invention relates to an α-amylase mutant of a parent α-amylase, comprising modifications at one or more positions corresponding to positions 3, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475, and 476, wherein the mutant has α-amylase activity.

[0171] In one embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 12 3, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 1 89, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, The variant includes modifications at one or more positions corresponding to positions 360, 361, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475, and 476, wherein the variant has α-amylase activity.

[0172] In one embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 1 60, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 361 , comprising modifications at one or more positions corresponding to positions 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0173] In one embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 16 7, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 361, 362, 365, 371, 383, 3 The variant includes modifications at one or more positions corresponding to positions 90, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, and at least Having sequence homology of 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%, the α-amylase variant has improved properties compared to the parent polypeptide.

[0174] In one embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128 ,131,132,134,135,136,137,138,140,142,144,146,151,152,155,159,160,165,167,169,172,173,174,176,177,178,180,181,182,183,185,188,189,193,194,196,202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 361, 362, 365, 371, 383, 390, The variant includes modifications at one or more positions corresponding to positions 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475, and 476, wherein the variant has α-amylase activity, and the α-amylase variant has improved properties compared to the parent polypeptide.

[0175] In one embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 1 32, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 361, 362, 365, 371, 383, 390, 394, 395, 405, 409, 41 The variant includes modifications at one or more positions corresponding to positions 4, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475, and 476, wherein the variant has α-amylase activity, the α-amylase variant has improved properties compared to the parent polypeptide, and the parent polypeptide is SEQ ID NO: 1, 2, or 3.

[0176] In another embodiment, the variant has sequence homology with the parent polypeptide of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0177] In another embodiment, the mutant has sequence homology with the parent polypeptide having α-amylase activity of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0178] In one embodiment, the variant has sequence homology with the hybrid polypeptide of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0179] In one embodiment, the mutant has sequence homology with a hybrid polypeptide having α-amylase activity of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0180] In another embodiment, the variant has sequence homology with the polypeptide of SEQ ID NO: 1 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0181] In another embodiment, the variant has sequence homology with the polypeptide of SEQ ID NO: 2 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0182] In another embodiment, the variant has sequence homology with the polypeptide of Sequence ID No. 3 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0183] In another embodiment, the variant has sequence homology with the polypeptide of SEQ ID NO: 4 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0184] In another embodiment, the variant has sequence homology with the polypeptide of Sequence ID No. 5 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0185] In another embodiment, the variant has sequence homology with the polypeptide of Sequence ID No. 6 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0186] In another embodiment, the variant has sequence homology with the polypeptide of Sequence ID No. 7 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0187] In another embodiment, the variant has sequence homology with the polypeptide of Sequence ID No. 8 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0188] In another embodiment, the variant has sequence homology with the polypeptide of Sequence ID No. 9 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0189] In another embodiment, the variant has sequence homology with the polypeptide of SEQ ID NO: 10 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0190] In another embodiment, the variant has sequence homology with the polypeptide of Sequence ID No. 11 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0191] In another embodiment, the variant has sequence homology with the polypeptide of SEQ ID NO: 12 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0192] In another embodiment, the variant has sequence homology with the polypeptide of Sequence ID No. 13 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0193] In another embodiment, the variant has sequence homology with the polypeptide of Sequence ID No. 14 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0194] In one embodiment, the substituted amino acid residue is different from the naturally occurring amino acid residue at that position. In one embodiment, the substitution is selected from the group consisting of A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W, and Y, provided that the substituted amino acid residue is different from the naturally occurring amino acid residue at that position.

[0195] In one embodiment, the number of modifications is 1 to 50, for example, 1 to 45, 1 to 40, 1 to 35, 1 to 30, 1 to 25, 1 to 20, 1 to 15, 1 to 10, or 1 to 5, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50 modifications.

[0196] In one embodiment, the number of substitutions is 1 to 50, for example, 1 to 45, 1 to 40, 1 to 35, 1 to 30, 1 to 25, 1 to 20, 1 to 15, 1 to 10, or 1 to 5, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50 substitutions.

[0197] In one embodiment, the number of deletions is 1 to 50, for example, 1 to 45, 1 to 40, 1 to 35, 1 to 30, 1 to 25, 1 to 20, 1 to 15, 1 to 10, or 1 to 5, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50 deletions.

[0198] The aforementioned variants with substitutions should be understood to encompass all possible combinations of one or more substitutions at a particular location.

[0199] In one embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 1 60, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 361 , comprising modifications at one or more positions corresponding to positions 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0200] In one embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160 ,165,167,169,172,173,174,176,177,178,180,181,182,183,185,188,189,193,194,196,202,203,205,224,225,242,243,244,252,256,258,259,260,261,266,276,279,280,283,286,302,303,315,320,321,322,323,346,354,359,360,361,362, The variant includes modifications at one or more (e.g., several) positions corresponding to positions 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, less than 14 of SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 or 14 At least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but with less than 100% sequence homology.

[0201] In another embodiment, the α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159 ,160,165,167,169,172,173,174,176,177,178,180,181,182,183,185,188,189,193,194,196,202,203,205,224,225,242,243,244,252,256,258,259,260,261,266,276,279,280,283,286,302,303,315,320,321,322,323,346,354,359,360,3 The variant includes modifications at two positions corresponding to positions 61, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0202] In another embodiment, the α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159 ,160,165,167,169,172,173,174,176,177,178,180,181,182,183,185,188,189,193,194,196,202,203,205,224,225,242,243,244,252,256,258,259,260,261,266,276,279,280,283,286,302,303,315,320,321,322,323,346,354,359,360,3 The variant includes modifications at three positions corresponding to positions 61, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0203] In another embodiment, the α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159 ,160,165,167,169,172,173,174,176,177,178,180,181,182,183,185,188,189,193,194,196,202,203,205,224,225,242,243,244,252,256,258,259,260,261,266,276,279,280,283,286,302,303,315,320,321,322,323,346,354,359,360,3 The variant includes modifications at four positions corresponding to positions 61, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0204] In another embodiment, the α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159 ,160,165,167,169,172,173,174,176,177,178,180,181,182,183,185,188,189,193,194,196,202,203,205,224,225,242,243,244,252,256,258,259,260,261,266,276,279,280,283,286,302,303,315,320,321,322,323,346,354,359,360,3 The variant includes modifications at five positions corresponding to positions 61, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0205] In another embodiment, the α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159 ,160,165,167,169,172,173,174,176,177,178,180,181,182,183,185,188,189,193,194,196,202,203,205,224,225,242,243,244,252,256,258,259,260,261,266,276,279,280,283,286,302,303,315,320,321,322,323,346,354,359,360,3 The variant includes modifications at six positions corresponding to positions 61, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0206] In another embodiment, the α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159 ,160,165,167,169,172,173,174,176,177,178,180,181,182,183,185,188,189,193,194,196,202,203,205,224,225,242,243,244,252,256,258,259,260,261,266,276,279,280,283,286,302,303,315,320,321,322,323,346,354,359,360,3 The variant includes modifications at seven positions corresponding to positions 61, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0207] In another embodiment, the α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159 ,160,165,167,169,172,173,174,176,177,178,180,181,182,183,185,188,189,193,194,196,202,203,205,224,225,242,243,244,252,256,258,259,260,261,266,276,279,280,283,286,302,303,315,320,321,322,323,346,354,359,360,3 The variant includes modifications at eight positions corresponding to positions 61, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0208] In another embodiment, the α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159 ,160,165,167,169,172,173,174,176,177,178,180,181,182,183,185,188,189,193,194,196,202,203,205,224,225,242,243,244,252,256,258,259,260,261,266,276,279,280,283,286,302,303,315,320,321,322,323,346,354,359,360,3 The variant includes modifications at nine positions corresponding to positions 61, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0209] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 10 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0210] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 11 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0211] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 12 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0212] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 13 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0213] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 14 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0214] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 15 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0215] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 16 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0216] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 17 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0217] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 18 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0218] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 19 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0219] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 20 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0220] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 21 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0221] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 22 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0222] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 23 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0223] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 24 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0224] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 25 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0225] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 26 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0226] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at 27 positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having less than 100% sequence homology.

[0227] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 167, 169, 172, 173, 174, 176, 177, 178, 180, 181, 182, 183, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 36 The variant includes modifications at the positions corresponding to positions 1, 362, 365, 371, 383, 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, and at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, having sequence homology of less than 100%.

[0228] In another embodiment, the mutant contains or consists of a deletion at the position corresponding to position 1. In another embodiment, the amino acid at the position corresponding to position 1 of the polypeptide of SEQ ID NO: 1 is deleted. In another embodiment, the mutant contains or consists of a deletion H1* of the polypeptide of SEQ ID NO: 1.

[0229] In another embodiment, the mutant contains or consists of a deletion at the position corresponding to position 2. In another embodiment, the amino acid at the position corresponding to position 2 of the polypeptide of SEQ ID NO: 1 is deleted. In another embodiment, the mutant contains or consists of a deletion H2* of the polypeptide of SEQ ID NO: 1.

[0230] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 3 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 3 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution N3S, N3D, or N3G of the polypeptide of SEQ ID NO: 1.

[0231] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 7 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 7 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution G7A, G7H, or G7S of the polypeptide of SEQ ID NO: 1.

[0232] In another embodiment, the mutant contains or consists of a substitution at the position corresponding to position 26 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 26 is substituted with Ala, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the mutant contains or consists of the substitution R26Y of the polypeptide of SEQ ID NO: 1.

[0233] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 31 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 31 is substituted with Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution A31S of the polypeptide of SEQ ID NO: 1.

[0234] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 37 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 37 is substituted with Ala, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the polypeptide substitution R37A, R37N, or R37V of SEQ ID NO: 1.

[0235] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 40 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 40 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the T40N or T40G substitution of the polypeptide of SEQ ID NO: 1.

[0236] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 41 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 41 is substituted with Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Trp, Thr, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution A41D of the polypeptide of SEQ ID NO: 1.

[0237] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 54 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 54 is substituted with Ala, Arg, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution N54S of the polypeptide of SEQ ID NO: 1.

[0238] In another embodiment, the mutant contains or consists of a substitution at the position corresponding to position 56 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 56 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, or Tyr. In yet another embodiment, the mutant contains or consists of the substitution V56T of the polypeptide of SEQ ID NO: 1.

[0239] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 60 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 60 is substituted with Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the polypeptide substitution A60P or A60V of SEQ ID NO: 1.

[0240] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 63 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 63 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution L63F of the polypeptide of SEQ ID NO: 1.

[0241] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 72 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 72 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution K72R of the polypeptide of SEQ ID NO: 1.

[0242] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 73 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 73 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution G73L of the polypeptide of SEQ ID NO: 1.

[0243] In another embodiment, the mutant contains or consists of a substitution at the position corresponding to position 93 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 93 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the mutant contains or consists of the substitution K93D of the polypeptide of SEQ ID NO: 1.

[0244] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 94 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 94 is substituted with Ala, Arg, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution N94D of the polypeptide of SEQ ID NO: 1.

[0245] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 98 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 98 is substituted with Ala, Arg, Asn, Asp, Cys, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution Q98L of the polypeptide of SEQ ID NO: 1.

[0246] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 103 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 103 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, or Tyr. In yet another embodiment, the variant contains or consists of the substitution V103 or VA103I of the polypeptide of SEQ ID NO: 1.

[0247] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 104 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 104 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, or Tyr. In yet another embodiment, the variant contains or consists of the substitution V104M of the polypeptide of SEQ ID NO: 1.

[0248] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 105 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 105 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the polypeptide substitution M105I or M105L of SEQ ID NO: 1.

[0249] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 106 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 106 is substituted with Ala, Arg, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution N106D of the polypeptide of SEQ ID NO: 1.

[0250] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 109 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 109 is substituted with Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution A109G of the polypeptide of SEQ ID NO: 1.

[0251] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 110 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 110 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution G110K of the polypeptide of SEQ ID NO: 1.

[0252] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 111 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 111 is substituted with Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the polypeptide substitution A111G or A111L of SEQ ID NO: 1.

[0253] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 113 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 113 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the polypeptide substitution G113A, G113Q, or G113S of SEQ ID NO: 1.

[0254] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 114 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 114 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the T114K substitution of the polypeptide of SEQ ID NO: 1.

[0255] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 116 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 116 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the polypeptide substitution F116L, F116N, F116Q, or F116W of SEQ ID NO: 1.

[0256] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 117 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 117 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, or Tyr. In yet another embodiment, the variant contains or consists of the substitution V117G of the polypeptide of SEQ ID NO: 1.

[0257] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 118 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 118 is substituted with Ala, Arg, Asn, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the D118T or D118Q substitution of the polypeptide of SEQ ID NO: 1.

[0258] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 123 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 123 is substituted with Ala, Arg, Asn, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution D123N of the polypeptide of SEQ ID NO: 1.

[0259] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 126 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 126 is substituted with Ala, Arg, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution N126D of the polypeptide of SEQ ID NO: 1.

[0260] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 128 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 128 is substituted with Ala, Arg, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution N128Y of the polypeptide of SEQ ID NO: 1.

[0261] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 131 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 131 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the T131I or T131V substitution of the polypeptide of SEQ ID NO: 1.

[0262] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 132 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 132 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution S132D or S132L of the polypeptide of SEQ ID NO: 1.

[0263] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 134 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 134 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution T134D of the polypeptide of SEQ ID NO: 1.

[0264] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 135 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 135 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, or Val. In yet another embodiment, the variant contains or consists of the substitution Y135E of the polypeptide of SEQ ID NO: 1.

[0265] In another embodiment, the mutant contains or consists of a substitution at the position corresponding to position 136 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 136 is substituted with Ala, Arg, Asn, Asp, Cys, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the mutant contains or consists of the substitution Q136T or Q136K of the polypeptide of SEQ ID NO: 1.

[0266] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 137 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 137 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution I137F of the polypeptide of SEQ ID NO: 1.

[0267] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 138 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 138 is substituted with Ala, Arg, Asn, Asp, Cys, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution Q138V of the polypeptide of SEQ ID NO: 1.

[0268] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 140 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 140 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Tyr, or Val. In another embodiment, the variant contains or consists of the polypeptide substitution W140S, W140Y, or W140M of SEQ ID NO: 1.

[0269] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 142 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 142 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution K142E, K142G, or K142P of the polypeptide of SEQ ID NO: 1.

[0270] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 144 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 144 is substituted with Ala, Arg, Asn, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution D144N of the polypeptide of SEQ ID NO: 1.

[0271] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 146 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 146 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution P146H or P146S of the polypeptide of SEQ ID NO: 1.

[0272] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 151 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 151 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the T151A substitution of the polypeptide of SEQ ID NO: 1.

[0273] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 152 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 152 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, or Val. In yet another embodiment, the variant contains or consists of the substitution Y152L of the polypeptide of SEQ ID NO: 1.

[0274] In another embodiment, the mutant contains or consists of a substitution at the position corresponding to position 155 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 155 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the mutant contains or consists of the substitution F155W of the polypeptide of SEQ ID NO: 1.

[0275] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 159 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 159 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Tyr, or Val. In another embodiment, the variant contains or consists of the W159H or W159L substitution of the polypeptide of SEQ ID NO: 1.

[0276] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 160 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 160 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, or Val. In another embodiment, the variant contains or consists of the substitution Y160P of the polypeptide of SEQ ID NO: 1.

[0277] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 165 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 165 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution T165V of the polypeptide of SEQ ID NO: 1.

[0278] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 167 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 167 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Tyr, or Val. In another embodiment, the variant contains or consists of the polypeptide substitution W167A, W167D, W167E, W167F, W167P, or W167S of SEQ ID NO: 1.

[0279] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 169 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 169 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Gly, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the polypeptide substitution E169V, E169I, or E169Q of SEQ ID NO: 1.

[0280] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 172 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 172 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the K172G or K172Q substitution of the polypeptide of SEQ ID NO: 1.

[0281] In another aspect, the variant comprises or consists of a substitution at a position corresponding to position 173 of SEQ ID NO: 1. In another aspect, the amino acid at the position corresponding to position 173 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another aspect, the variant comprises or consists of the substitution L173F of the polypeptide of SEQ ID NO: 1.

[0282] In another aspect, the variant comprises or consists of a substitution at a position corresponding to position 174 of SEQ ID NO: 1. In another aspect, the amino acid at the position corresponding to position 174 is substituted with Ala, Arg, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another aspect, the variant comprises or consists of the substitution N174K or N174S of the polypeptide of SEQ ID NO: 1.

[0283] In another aspect, the variant comprises or consists of a substitution at a position corresponding to position 176 of SEQ ID NO: 1. In another aspect, the amino acid at the position corresponding to position 176 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another aspect, the variant comprises or consists of the substitution I176V of the polypeptide of SEQ ID NO: 1.

[0284] In another aspect, the variant comprises or consists of a substitution at a position corresponding to position 177 of SEQ ID NO: 1. In another aspect, the amino acid at the position corresponding to position 177 is substituted with Ala, Arg, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, or Val. In another aspect, the variant comprises or consists of the substitution Y177D of the polypeptide of SEQ ID NO: 1.

[0285] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 178 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 178 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution K178A of the polypeptide of SEQ ID NO: 1.

[0286] In another embodiment, the mutant contains or consists of a deletion or substitution at the position corresponding to position 180. In another embodiment, the amino acid at the position corresponding to position 180 of the polypeptide of SEQ ID NO: 1 is deleted. In another embodiment, the mutant contains or consists of the deletion R180* of the polypeptide of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 180 is substituted with Ala, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the mutant contains or consists of the substitution R180Q or R180S of the polypeptide of SEQ ID NO: 1.

[0287] In another embodiment, the mutant contains or consists of a deletion or substitution at the position corresponding to position 181. In another embodiment, the amino acid at the position corresponding to position 181 of the polypeptide of SEQ ID NO: 1 is deleted. In another embodiment, the mutant contains or consists of the deletion S181* of the polypeptide of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 181 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Thr, Trp, Tyr, or Val. In another embodiment, the mutant contains or consists of the substitution S181L or S181T of the polypeptide of SEQ ID NO: 1.

[0288] In another embodiment, the mutant includes or consists of a deletion or substitution at the position corresponding to position 182 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 182 of the polypeptide of SEQ ID NO: 1 is deleted. In another embodiment, the mutant includes or consists of the deletion T182* of the polypeptide of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 182 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Trp, Tyr, or Val. In another embodiment, the mutant includes or consists of the substitution T182D, T182E, T182G, T182I, T182L, T182M, or T182S of the polypeptide of SEQ ID NO: 1.

[0289] In another embodiment, the mutant contains or consists of a deletion or substitution at the position corresponding to position 183 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 183 of the polypeptide of SEQ ID NO: 1 is deleted. In another embodiment, the mutant contains or consists of the deletion G183* of the polypeptide of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 183 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the mutant contains or consists of the substitution G183P of the polypeptide of SEQ ID NO: 1.

[0290] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 185 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 185 is substituted with Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the polypeptide substitution A186D, A186E, A186K, A186N, A186Q, or A186R of SEQ ID NO: 1.

[0291] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 188 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 188 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution W188C of the polypeptide of SEQ ID NO: 1.

[0292] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 189 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 189 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution E189P of the polypeptide of SEQ ID NO: 1.

[0293] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 193 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 193 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution E193I of the polypeptide of SEQ ID NO: 1.

[0294] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 194 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 194 is substituted with Ala, Arg, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution N194F of the polypeptide of SEQ ID NO: 1.

[0295] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 196 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 196 is substituted with Ala, Arg, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution N196F or N196I of the polypeptide of SEQ ID NO: 1.

[0296] In another embodiment, the mutant contains or consists of a substitution at the position corresponding to position 202 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 202 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the mutant contains or consists of the substitution F202Y of the polypeptide of SEQ ID NO: 1.

[0297] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 203 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 203 is substituted with Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution A203F of the polypeptide of SEQ ID NO: 1.

[0298] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 205 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 205 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the polypeptide substitution L205F, L205N, L205Q, L205R, L205V, or L205Y of SEQ ID NO: 1.

[0299] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 224 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 224 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the polypeptide substitution T224F or T244Y of SEQ ID NO: 1.

[0300] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 225 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 225 is substituted with Ala, Arg, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In yet another embodiment, the variant contains or consists of the substitution N225S of the polypeptide of SEQ ID NO: 1.

[0301] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 242 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 242 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, or Val. In another embodiment, the variant contains or consists of the substitution Y242F or Y242L of the polypeptide of SEQ ID NO: 1.

[0302] In another embodiment, the mutant contains or consists of a substitution at the position corresponding to position 243 of SEQ ID NO: 1. In another embodiment, the amino acid at the position corresponding to position 243 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Thr, Trp, Tyr, or Val. In yet another embodiment, the mutant contains or consists of the substitution S243Q of the polypeptide of SEQ ID NO: 1.

[0303] In another aspect, the variant comprises or consists of a substitution at the position corresponding to position 244 of SEQ ID NO: 1. In another aspect, the amino acid at the position corresponding to position 244 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Ser, Pro, Thr, Trp, Tyr, or Val. In another aspect, the variant comprises or consists of the substitution F244I or F244L of the polypeptide of SEQ ID NO: 1.

[0304] In another aspect, the variant comprises or consists of a substitution at the position corresponding to position 252 of SEQ ID NO: 1. In another aspect, the amino acid at the position corresponding to position 252 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, or Tyr. In another aspect, the variant comprises or consists of the substitution V252I of the polypeptide of SEQ ID NO: 1.

[0305] In another aspect, the variant comprises or consists of a substitution at the position corresponding to position 256 of SEQ ID NO: 1. In another aspect, the amino acid at the position corresponding to position 256 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Trp, Tyr, or Val. In another aspect, the variant comprises or consists of the substitution T256A or T256D of the polypeptide of SEQ ID NO: 1.

[0306] In another aspect, the variant comprises or consists of a substitution at the position corresponding to position 258. In another aspect, the amino acid at the position corresponding to position 258 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another aspect, the variant comprises or consists of the substitution K258V of the polypeptide of SEQ ID NO: 1.

[0307] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 259. In another embodiment, the amino acid at the position corresponding to position 259 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution E259G or E259V of the polypeptide of SEQ ID NO: 1.

[0308] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 260. In another embodiment, the amino acid at the position corresponding to position 260 is substituted with Ala, Arg, Asn, Asp, Cys, Glu, Gln, Gly, His, Ile, Leu, Lys, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the polypeptide substitution M260D or M260W of SEQ ID NO: 1.

[0309] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 261. In another embodiment, the amino acid at the position corresponding to position 261 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the polypeptide substitution F261K, F261P, or F261S of SEQ ID NO: 1.

[0310] In another embodiment, the mutant contains or consists of a substitution at the position corresponding to position 266. In another embodiment, the amino acid at the position corresponding to position 266 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the mutant contains or consists of the substitution F266Y of the polypeptide of SEQ ID NO: 1.

[0311] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 276. In another embodiment, the amino acid at the position corresponding to position 276 is substituted with Ala, Arg, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution N276F, N276W, or N276Y of the polypeptide of SEQ ID NO: 1.

[0312] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 279. In another embodiment, the amino acid at the position corresponding to position 279 is substituted with Ala, Arg, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution N279S of the polypeptide of SEQ ID NO: 1.

[0313] In another embodiment, the mutant contains or consists of a substitution at the position corresponding to position 280. In another embodiment, the amino acid at the position corresponding to position 280 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the mutant contains or consists of the polypeptide substitution K280F, K280W, or K280Y of SEQ ID NO: 1.

[0314] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 283. In another embodiment, the amino acid at the position corresponding to position 283 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution W283N of the polypeptide of SEQ ID NO: 1.

[0315] In another embodiment, the mutant contains or consists of a substitution at the position corresponding to position 286. In another embodiment, the amino acid at the position corresponding to position 286 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Thr, Trp, Tyr, or Val. In another embodiment, the mutant contains or consists of the substitution S286K or S286L of the polypeptide of SEQ ID NO: 1.

[0316] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 302. In another embodiment, the amino acid at the position corresponding to position 302 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution S302T or S302Y of the polypeptide of SEQ ID NO: 1.

[0317] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 303. In another embodiment, the amino acid at the position corresponding to position 303 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution G303R of the polypeptide of SEQ ID NO: 1.

[0318] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 315. In another embodiment, the amino acid at the position corresponding to position 315 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the T315A substitution of the polypeptide of SEQ ID NO: 1.

[0319] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 320. In another embodiment, the amino acid at the position corresponding to position 320 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the H320T substitution of the polypeptide of SEQ ID NO: 1.

[0320] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 321. In another embodiment, the amino acid at the position corresponding to position 321 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution P321H or P321Y of the polypeptide of SEQ ID NO: 1.

[0321] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 322. In another embodiment, the amino acid at the position corresponding to position 322 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the polypeptide substitution M322G or M322Y of SEQ ID NO: 1.

[0322] In another embodiment, the mutant contains or consists of a substitution at the position corresponding to position 323. In another embodiment, the amino acid at the position corresponding to position 323 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the mutant contains or consists of the H323A, H323N, or H323W substitution of the polypeptide of SEQ ID NO: 1.

[0323] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 346. In another embodiment, the amino acid at the position corresponding to position 346 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution W346N of the polypeptide of SEQ ID NO: 1.

[0324] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 354. In another embodiment, the amino acid at the position corresponding to position 354 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution L354V of the polypeptide of SEQ ID NO: 1.

[0325] In another embodiment, the mutant contains or consists of a substitution at the position corresponding to position 359. In another embodiment, the amino acid at the position corresponding to position 359 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the mutant contains or consists of the substitution E359W of the polypeptide of SEQ ID NO: 1.

[0326] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 360. In another embodiment, the amino acid at the position corresponding to position 360 is substituted with Ala, Arg, Asn, Asp, Cys, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution Q360M, Q360W, or Q360Y of the polypeptide of SEQ ID NO: 1.

[0327] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 361. In another embodiment, the amino acid at the position corresponding to position 361 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, or Tyr. In another embodiment, the variant contains or consists of the substitution G361M of the polypeptide of SEQ ID NO: 1.

[0328] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 362. In another embodiment, the amino acid at the position corresponding to position 362 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, or Val. In another embodiment, the variant contains or consists of the substitution Y362V of polypeptide SEQ ID NO: 1.

[0329] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 365. In another embodiment, the amino acid at the position corresponding to position 365 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Tyr, or Trp. In another embodiment, the variant contains or consists of the substitution V365I of the polypeptide of SEQ ID NO: 1.

[0330] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 371. In another embodiment, the amino acid at the position corresponding to position 371 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, or Val. In another embodiment, the variant contains or consists of the substitution Y371S or Y371N of the polypeptide of SEQ ID NO: 1.

[0331] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 383. In another embodiment, the amino acid at the position corresponding to position 383 is substituted with Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution A383D of the polypeptide of SEQ ID NO: 1.

[0332] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 390. In another embodiment, the amino acid at the position corresponding to position 390 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution E390A or E390D of the polypeptide of SEQ ID NO: 1.

[0333] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 394. In another embodiment, the amino acid at the position corresponding to position 394 is substituted with Ala, Arg, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution N394W or N394D of the polypeptide of SEQ ID NO: 1.

[0334] In another embodiment, the mutant contains or consists of a substitution at the position corresponding to position 395. In another embodiment, the amino acid at the position corresponding to position 395 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the mutant contains or consists of the substitution F395Y of the polypeptide of SEQ ID NO: 1.

[0335] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 405. In another embodiment, the amino acid at the position corresponding to position 405 is substituted with Ala, Arg, Asn, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the D405G substitution of the polypeptide of SEQ ID NO: 1.

[0336] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 409. In another embodiment, the amino acid at the position corresponding to position 409 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution I409L of the polypeptide of SEQ ID NO: 1.

[0337] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 414. In another embodiment, the amino acid at the position corresponding to position 414 is substituted with Ala, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution R414M of the polypeptide of SEQ ID NO: 1.

[0338] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 420. In another embodiment, the amino acid at the position corresponding to position 420 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the polypeptide substitution H420A, H420P, H420Q, or H420Y of SEQ ID NO: 1.

[0339] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 422. In another embodiment, the amino acid at the position corresponding to position 422 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, or Tyr. In another embodiment, the variant contains or consists of the substitution N422V of the polypeptide of SEQ ID NO: 1.

[0340] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 427. In another embodiment, the amino acid at the position corresponding to position 427 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the T427L substitution of the polypeptide of SEQ ID NO: 1.

[0341] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 455. In another embodiment, the amino acid at the position corresponding to position 455 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution G455R of the polypeptide of SEQ ID NO: 1.

[0342] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 457. In another embodiment, the amino acid at the position corresponding to position 457 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution K457N or K457R of the polypeptide of SEQ ID NO: 1.

[0343] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 458. In another embodiment, the amino acid at the position corresponding to position 458 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution P458S of the polypeptide of SEQ ID NO: 1.

[0344] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 470. In another embodiment, the amino acid at the position corresponding to position 470 is substituted with Ala, Arg, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution N470A or N470M of the polypeptide of SEQ ID NO: 1.

[0345] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 472. In another embodiment, the amino acid at the position corresponding to position 472 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution S472R of the polypeptide of SEQ ID NO: 1.

[0346] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 473. In another embodiment, the amino acid at the position corresponding to position 473 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, or Tyr. In another embodiment, the variant contains or consists of the substitution V473C of the polypeptide of SEQ ID NO: 1.

[0347] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 474. In another embodiment, the amino acid at the position corresponding to position 474 is substituted with Ala, Arg, Asp, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution N474F or N474L of the polypeptide of SEQ ID NO: 1.

[0348] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 475. In another embodiment, the amino acid at the position corresponding to position 475 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, His, Ile, Leu, Lys, Met, Pro, Phe, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the substitution G475E or G475K of the polypeptide of SEQ ID NO: 1.

[0349] In another embodiment, the variant contains or consists of a substitution at the position corresponding to position 476. In another embodiment, the amino acid at the position corresponding to position 476 is substituted with Ala, Arg, Asn, Asp, Cys, Gln, Glu, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, or Val. In another embodiment, the variant contains or consists of the polypeptide substitution G476A, G476E, or G476H of SEQ ID NO: 1.

[0350] In another embodiment, α-amylase mutants are numbered using SEQ ID NO: 1, H1, H2, N3, G7, R26, A31, R37, T40, A41, N54, V56, A60, L63, K72, G73, K93, N94, Q98, V103, V104, M105, N106, A109, G110, A111, G113, T114, F116, V117, D118, D123, N126, N128, T131, S132, T134, Y135, Q136, I137, Q138, W140, K142, D144, P146, T1 51, Y152, F155, W159, Y160, T165, W167, E169, K172, L173, N174, I176, Y 177, K178, R180, S181, T182, G183, A185, W188, E189, E193, N194, N196, F202, A203, L205, T224, N225, Y242, S243, F244, V252, T256, K258, E259 , M260, F261, F266, N276, N279, K280, W283, S286, S302, G303, T315, H320 , comprising modifications at one or more positions corresponding to positions P321, M322, H323, W346, L354, E359, Q360, G361, Y362, V365, Y371, A383, E390, N394, F395, D405, I409, R414, H420, N422, T427, G455, K457, P458, N470, S472, V473, N474, G475 and G476, wherein the variant has α-amylase activity, and the variant is less than 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 or 14. At least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but with less than 100% sequence homology.

[0351] In another embodiment, α-amylase variants are numbered using SEQ ID NO: 1, as follows: H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F 116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T13 1I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F1 55W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S , E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K 178A, R180*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T18 2G, T182I, T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N , A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L2 05N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243 Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F 261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N28 0Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y,M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361M, Y3 62V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F395Y, D405G, I409L, R414M The modification includes one or more positions corresponding to H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E and G476A, and the said modification The variants possess α-amylase activity, and the variants have sequence homology with SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0352] In another embodiment, α-amylase variants are numbered using SEQ ID NO: 1, as follows: H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F 116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T13 1I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F1 55W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S , E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K 178A, R180*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T18 2G, T182I, T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N , A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L2 05N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243 Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F 261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N28 0Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y,M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361 M, Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F395Y, D405G, I40 1 corresponding to 9L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E and G476A The variant comprises modifications at one or more positions, the variant has α-amylase activity, and the variant has sequence homology with SEQ ID NO: 1 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0353] In another embodiment, α-amylase variants are numbered using SEQ ID NO: 1, as follows: H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F 116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T13 1I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F1 55W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S , E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K 178A, R180*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T18 2G, T182I, T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N , A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L2 05N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243 Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F 261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N28 0Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y,M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361 M, Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F395Y, D405G, I40 1 corresponding to 9L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E and G476A The variant comprises modifications at one or more positions, the variant has α-amylase activity, and the variant has sequence homology with SEQ ID NO: 2 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0354] In another embodiment, α-amylase variants are numbered using SEQ ID NO: 1, as follows: H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F 116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T13 1I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F1 55W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S , E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K 178A, R180*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T18 2G, T182I, T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N , A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L2 05N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243 Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F 261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N28 0Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y,M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361 M, Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F395Y, D405G, I40 1 corresponding to 9L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E and G476A The variant comprises modifications at one or more positions, the variant has α-amylase activity, and the variant has sequence homology with SEQ ID NO: 3 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0355] In another embodiment, α-amylase variants are numbered using SEQ ID NO: 1, as follows: H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F 116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T13 1I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F1 55W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S , E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K 178A, R180*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T18 2G, T182I, T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N , A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L2 05N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243 Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F 261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N28 0Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y,M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361 M, Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F395Y, D405G, I40 1 corresponding to 9L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E and G476A The variant includes modifications at one or more positions, the variant has α-amylase activity, and the variant has sequence homology with SEQ ID NO: 4 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0356] In another embodiment, α-amylase variants are numbered using SEQ ID NO: 1, as follows: H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F 116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T13 1I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F1 55W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S , E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K 178A, R180*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T18 2G, T182I, T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N , A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L2 05N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243 Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F 261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N28 0Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y,M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361 M, Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F395Y, D405G, I40 1 corresponding to 9L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E and G476A The variant comprises modifications at one or more positions, the variant has α-amylase activity, and the variant has sequence homology with SEQ ID NO: at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0357] In another embodiment, α-amylase variants are numbered using SEQ ID NO: 1, as follows: H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F 116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T13 1I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F1 55W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S , E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K 178A, R180*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T18 2G, T182I, T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N , A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L2 05N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243 Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F 261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N28 0Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y,M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361 M, Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F395Y, D405G, I40 1 corresponding to 9L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E and G476A The variant comprises modifications at one or more positions, the variant has α-amylase activity, and the variant has sequence homology with SEQ ID NO: 6 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0358] In another embodiment, α-amylase variants are numbered using SEQ ID NO: 1, as follows: H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F 116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T13 1I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F1 55W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S , E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K 178A, R180*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T18 2G, T182I, T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N , A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L2 05N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243 Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F 261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N28 0Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y,M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361 M, Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F395Y, D405G, I40 1 corresponding to 9L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E and G476A The variant comprises modifications at one or more positions, the variant has α-amylase activity, and the variant has sequence homology with SEQ ID NO: 7 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0359] In another embodiment, α-amylase variants are numbered using SEQ ID NO: 1, as follows: H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F 116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T13 1I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F1 55W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S , E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K 178A, R180*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T18 2G, T182I, T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N , A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L2 05N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243 Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F 261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N28 0Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y,M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361 M, Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F395Y, D405G, I40 1 corresponding to 9L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E and G476A The variant includes modifications at one or more positions, the variant has α-amylase activity, and the variant has sequence homology with SEQ ID NO: 8 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0360] In another embodiment, α-amylase variants are numbered using SEQ ID NO: 1, as follows: H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F 116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T13 1I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F1 55W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S , E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K 178A, R180*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T18 2G, T182I, T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N , A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L2 05N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243 Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F 261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N28 0Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y,M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361 M, Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F395Y, D405G, I40 1 corresponding to 9L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E and G476A The variant comprises modifications at one or more positions, the variant has α-amylase activity, and the variant has sequence homology with SEQ ID NO: 9 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0361] In another embodiment, α-amylase variants are numbered using SEQ ID NO: 1, as follows: H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F 116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T13 1I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F1 55W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S , E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K 178A, R180*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T18 2G, T182I, T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N , A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L2 05N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243 Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F 261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N28 0Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y,M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361M , Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F395Y, D405G, I409 One corresponding to L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E, and G476A. The variant includes modifications at the above positions, possesses α-amylase activity, and has sequence homology with SEQ ID NO: 10 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0362] In another embodiment, α-amylase variants are numbered using SEQ ID NO: 1, as follows: H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F 116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T13 1I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F1 55W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S , E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K 178A, R180*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T18 2G, T182I, T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N , A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L2 05N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243 Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F 261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N28 0Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y,M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361M , Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F395Y, D405G, I409 One corresponding to L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E, and G476A. The variant includes modifications at the above positions, possesses α-amylase activity, and has sequence homology with SEQ ID NO: 11 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0363] In another embodiment, α-amylase variants are numbered using SEQ ID NO: 1, as follows: H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F 116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T13 1I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F1 55W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S , E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K 178A, R180*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T18 2G, T182I, T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N , A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L2 05N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243 Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F 261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N28 0Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y,M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361M , Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F395Y, D405G, I409 One corresponding to L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E, and G476A. The variant includes modifications at the above positions, possesses α-amylase activity, and has sequence homology with SEQ ID NO: 12 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0364] In another embodiment, α-amylase variants are numbered using SEQ ID NO: 1, as follows: H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F 116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T13 1I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F1 55W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S , E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K 178A, R180*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T18 2G, T182I, T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N , A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L2 05N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243 Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F 261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N28 0Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y,M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361M , Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F395Y, D405G, I409 One corresponding to L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E, and G476A. The variant includes modifications at the above positions, possesses α-amylase activity, and has sequence homology with SEQ ID NO: 13 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0365] In another embodiment, α-amylase variants are numbered using SEQ ID NO: 1, as follows: H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F 116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T13 1I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F1 55W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S , E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K 178A, R180*, R180Q, R180S, S181*, S181L, S181T, T182*, T182D, T182E, T18 2G, T182I, T182L, T182M, T182S, G183*, G183P, A185D, A185E, A185K, A185N , A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A203F, L205F, L2 05N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F, Y242L, S243 Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M260D, M260W, F 261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280F, N280W, N28 0Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P321H, P321Y,M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q360M, Q360W, Q360Y, G361M , Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D, N394W, N394D, F395Y, D405G, I409 One corresponding to L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E, and G476A. The variant includes modifications at the above positions, possesses α-amylase activity, and has sequence homology with SEQ ID NO: 14 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0366] As used herein, the term “pairwise deletion” refers to the deletion of amino acids at two positions. These two positions may be adjacent to each other, but may be separated by one, two, three, four, or five amino acids.

[0367] In one embodiment, the mutant contains a pairwise deletion within the B domain of the parent polypeptide.

[0368] In one embodiment, the mutant contains a pairwise deletion within the B domain of the parent polypeptide having α-amylase activity.

[0369] As used herein, the term “pairwise deletion” refers to the deletion of amino acids at two positions. These two positions may be adjacent to each other, but may be separated by one, two, three, four, or five amino acids.

[0370] In one embodiment, the α-amylase mutant of the present invention contains a deletion at one or more positions selected from the group consisting of R180*+S181*, R180*+T182*, R180*+G183*, S181*+T182*, S181*+G183* and T182*+G183*, preferably R180*+S181*, using Sequence ID No. 1 for numbering.

[0371] In one embodiment, the present invention relates to a variant of Sequence ID No. 1, comprising a deletion at a position corresponding to R180*+S181* using Sequence ID No. 1 for numbering, wherein the variant has α-amylase activity, and the variant has sequence homology of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%.

[0372] Therefore, in one embodiment, the α-amylase variant is a) Using sequence number 1 for numbering, delete and / or substitute at two or more positions corresponding to positions 180, 181, 182 and 183, b) Using sequence number 1 for numbering: 1, 2, 3, 7, 26, 31, 37, 40, 41, 54, 56, 60, 63, 72, 73, 93, 94, 98, 103, 104, 105, 106, 109, 110, 111, 113, 114, 116, 117, 118, 123, 126, 128, 131, 132, 134, 135, 136, 137, 138, 140, 142, 144, 146, 151, 152, 155, 159, 160, 165, 1 67, 169, 172, 173, 174, 176, 177, 178, 185, 188, 189, 193, 194, 196, 202, 203, 205, 224, 225, 242, 243, 244, 252, 256, 258, 259, 260, 261, 266, 276, 279, 280, 283, 286, 302, 303, 315, 320, 321, 322, 323, 346, 354, 359, 360, 361, 362, 365, 371, 383 , comprising modifications at one or more positions corresponding to positions 390, 394, 395, 405, 409, 414, 420, 422, 427, 455, 457, 458, 470, 472, 473, 474, 475 and 476, wherein the variant has α-amylase activity, and the variant is at least 60%, at least 65%, at least 70%, at least 75%, less than 60%, at least 65%, at least 75%, Both have 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but with less than 100% sequence homology.

[0373] In one embodiment, the α-amylase variant is a) Using sequence number 1 for numbering, delete and / or substitute at two or more positions corresponding to R180, S181, T182 and G183, b) Use array number 1 for numbering: H1, H2*, N3, G7, R26, A31, R37, T40, A41, N54, V56, A60, L63, K72, G73, K93, N94, Q98, V103, V104, M105, N106, A109, G110, A111, G113, T114, F116, V117, D118, D123, N126, N128, T131, S132, T134, Y135, Q136, I137, Q138, W140, K142, D144, P146, T151, Y152, F155, W159, Y160, T165, W167, E169, K172, L173, N174, I176, Y177, K178, A185, W188, E189, E193, N194, N196, F202, A203, L205, T224, N225 , Y242, S243, F244, V252, T256, K258, E259, M260, F261, F266, N276, N279, K280, W283, S286, S302, G303, T315, H320, P321, M322, H323, W34 The variant comprises modifications at one or more positions corresponding to positions 6, L354, E359, Q360, G361, Y362, V365, Y371, A383, E390, N394, F395, D405, I409, R414, H420, N422, T427, G455, K457, P458, N470, S472, V473, N474, G475 and G476, wherein the variant has α-amylase activity, and the variant is at least 60% less than SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 or 14. At least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but with less than 100% sequence homology.

[0374] In one embodiment, the α-amylase variant is a) Deletions and / or substitutions at two or more positions corresponding to positions R180, S181, T182 and G183, using sequence number 1 for numbering, b) For numbering, use sequence number 1 to assign H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T131I, T131V, S132D, S1 32L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E , K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F155W, W159H, W159L, Y 160P, T165V, W167A, W167D, W167E, W167F, W167P, W167S, E169I, E169Q, E169 V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K178A, A185D, A185E, A185K, A185N, A185Q, A185R, W188C, E189P, E193I, N194F, F202Y, F202Y, A20 3F, L205F, L205N, L205Q, L205R, L205V, L205Y, T224F, T224Y, N225S, Y242F , Y242L, S243Q, F244I, F244L, V252I, T256A, T256D, K258V, E259G, E259V, M2 60D, M260W, F261K, F261P, F261S, F266Y, N276F, N276W, N276Y, N279S, N280 F, N280W, N280Y, W283N, S286K, S286L, S302T, S302Y, G303R, T315A, H320T, P 321H, P321Y, M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E359W, Q36 0M, Q360W, Q360Y, G361M, Y362V, V365I, Y371S, Y371N, A383D, E390A, E390D,The variants include modifications at one or more positions corresponding to the N394W, N394D, F395Y, D405G, I409L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G476E and G476A positions, and the variants have α-amylase activity, and the variants are sequence numbers 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 or Sequence 14 has at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but with less than 100% sequence homology.

[0375] In one embodiment, the α-amylase mutant of SEQ ID NO: 1 is a) Using sequence number 1 for numbering, deletes are made at positions corresponding to R180* and S181*, b) Use array index 1 for numbering: H1*+D118T, H1*+W140Y, H1*+T182S, H1*+A185D, H1*+E189P, H1*+Y242F, H1*+E259G, H1*+F266Y, H1*+N279S, H1*+K280Y, H1*+G303R, H1*+E390A, H1*+G475K, H1*+A185Q, H1*+N194F, D118T+W140Y, D118T+T182S, D118T+A185D, D118T+E189P, D118T+Y242F, D118T+E259G, D118T+F2 66Y, D118T+N279S, D118T+K280Y, D118T+G303R, D118T+E390A, D118T+G475 K, D118T+A185Q, D118T+N194F, W140Y+T182S, W140Y+A185D, W140Y+E189P, W 140Y+Y242F, W140Y+E259G, W140Y+F266Y, W140Y+N279S, W140Y+K280Y, W14 0Y+G303R, W140Y+E390A, W140Y+G475K, W140Y+A185Q, W140Y+N194F, T182S+ A185D, T182S+E189P, T182S+Y242F, T182S+E259G, T182S+F266Y, T182S+N2 79S, T182S+K280Y, T182S+G303R, T182S+E390A, T182S+G475K, T182S+A185Q , T182S+N194F, A185D+E189P, A185D+Y242F, A185D+E259G, A185D+F266Y, A 185D+N279S, A185D+K280Y, A185D+G303R, A185D+E390A, A185D+G475K, A185 D, +A185Q, A185D, +N194F, E189P+Y242F, E189P+E259G, E189P+F266Y, E189 P+N279S, E189P+K280Y, E189P+G303R, E189P+E390A, E189P+G475K, E189P+A 185Q, E189P+N194F, Y242F+E259G, Y242F+F266Y, Y242F+N279S, Y242F+K28 0Y, Y242F+G303R, Y242F+E390A, Y242F+G475K, Y242F+A185Q, Y242F+N194F,E259G+F266Y, E259G+N279S, E259G+K280Y, E259G+G303R, E259G+E390A, E259G+G475K, E259G+A185Q, E259G+N19 4F, F266Y+N279S, F266Y+K280Y, F266Y+G303R, F266Y+E390A, F266Y+G475K, F266Y+A185Q, F266Y+N194F, N279+K 280Y, N279S+G303R, N279S+E390A, N279S+G475K, N279S+A185Q, N279S+N194F, K280Y+G303R, K280Y+E390A, K280 Y+G475K, K280Y+A185Q, K280Y+N194F, G303R+E390A, G303R+G475K, G303R+A185Q, G303R+N194F, E390A+G475K, E3 The variant includes modifications at positions corresponding to 90A+A185Q, E390A+N194F, G475K+A185Q, G475K+N194F, and A185Q+N194F, wherein the variant is at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, and at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, provided that the sequence homology is less than 100%, and the variant has α-amylase activity.

[0376] In one embodiment, the α-amylase mutant of SEQ ID NO: 1 is a) Using sequence number 1 for numbering, deletes are made at positions corresponding to R180* and S181*, b) Using array index 1 for numbering: H1*+D118T+W140Y, H1*+D118T+T182S, H1*+D118T+A185D, H1*+D118T+E189P, H1*+D118T+Y242F, H1*+D118T+E259G, H1*+D118T+F266Y, H1*+D118T+N279S, H1*+D118T+K280Y, H1*+D118T+G303R, H1*+D118T+E390A, H1*+D118T+G475K, H1*+D118T+A185Q, H1*+D118T+N194F, H1*+W 140Y+T182S, H1*+W140Y+A185D, H1*+W140Y+E189P, H1*+W140Y+Y242F, H1* +W140Y+E259G, H1*+W140Y+F266Y, H1*+W140Y+N279S, H1*+W140Y+K280Y, H 1*+W140Y+G303R, H1*+W140Y+E390A, H1*+W140Y+G475K, H1*+W140Y+A185Q , H1*+W140Y+N194F, H1*+T182S+A185D, H1*+T182S+E189P, H1*+T182S+Y242 F, H1*+T182S+E259G, H1*+T182S+F266Y, H1*+T182S+N279S, H1*+T182S+K2 80Y, H1*+T182S+G303R, H1*+T182S+E390A, H1*+T182S+G475K, H1*+T182S+ A185Q, H1*+T182S+N194F, H1*+A185D+E189P, H1*+A185D+Y242F, H1*+A185 D+E259G, H1*+A185D+F266Y, H1*+A185D+N279S, H1*+A185D+K280Y, H1*+A18 5D+G303R, H1*+A185D+E390A, H1*+A185D+G475K, H1*+A185D+A185Q, H1*+A 185D+N194F, H1*+E189P+Y242F, H1*+E189P+E259G, H1*+E189P+F266Y, H1* +E189P+N279S, H1*+E189P+K280Y, H1*+E189P+G303R, H1*+E189P+E390A, H 1*+E189P+G475K, H1*+E189P+A185Q, H1*+E189P+N194F, H1*+Y242F+E259G,H1*+Y242F+F266Y、H1*+Y242F+N279S、H1*+Y242F+K280Y、H1*+Y242F+G303R、H1*+Y242F+E390A、H1*+Y242F+G475K、H1*+Y242F+A185Q、H1*+Y242F+N194F、H1*+E259G+F266Y、H1*+E259G+N279S、H1*+E259G+K280Y、H1*+E259G+G303R、H1*+E259G+E390A、H1*+E259G+G475K、H1*+E259G+A185Q、H1*+E259G+N194F、H1*+F266Y+N279S、H1*+F266Y+K280Y、H1*+F266Y+G303R、H1*+F266Y+E390A、H1*+F266Y+G475K、H1*+F266Y+A185Q、H1*+F266Y+N194F、H1*+N279S+K280Y、H1*+N279S+G303R、H1*+N279S+E390A、H1*+N279S+G475K、H1*+N279S+A185Q、H1*+N279S+N194F、H1*+K280Y+G303R、H1*+K280Y+E390A、H1*+K280Y+G475K、H1*+K280Y+A185Q、H1*+K280Y+N194F、H1*+G303R+E390A、H1*+G303R+G475K、H1*+G303R+A185Q、H1*+G303R+N194F、H1*+E390A+G475K、H1*+E390A+A185Q、H1*+E390A+N194F、H1*+G475K+A185Q、H1*+G475K+N194F、H1*+A185Q+N194F、D118T+W140Y+T182S、D118T+W140Y+A185D、D118T+W140Y+E189P、D118T+W140Y+Y242F、D118T+W140Y+E259G、D118T+W140Y+F266Y、D118T+W140Y+N279S、D118T+W140Y+K280Y、D118T+W140Y+G303R、D118T+W140Y+E390A、D118T+W140Y+G475K、D118T+W140Y+A185Q、D118T+W140Y+N194F、D118T+T182S+A185D、D118T+T182S+E189P、D118T+T182S+Y242F、D118T+T182S+E259G、D118T+T182S+F266Y、D118T+T182S+N279S、D118T+T182S+K280Y、D118T+T182S+G303R、D118T+T182S+E390A、D118T+T182S+G475K、D118T+T182S+A185Q、D118T+T182S+N194F、D118T+A185D+E189P、D118T+A185D+Y242F、D118T+A185D+E259G、D118T+A185D+F266Y、D118T+A185D+N279S、D118T+A185D+K280Y、D118T+A185D+G303R、D118T+A185D+E390A、D118T+A185D+G475K、D118T+A185D+A185Q、D118T+A185D+N194F、D118T+E189P+Y242F、D118T+E189P+E259G、D118T+E189P+F266Y、D118T+E189P+N279S、D118T+E189P+K280Y、D118T+E189P+G303R、D118T+E189P+E390A、D118T+E189P+G475K、D118T+E189P+A185Q、D118T+E189P+N194F、D118T+Y242F+E259G、D118T+Y242F+F266Y、D118T+Y242F+N279S、D118T+Y242F+K280Y、D118T+Y242F+G303R、D118T+Y242F+E390A、D118T+Y242F+G475K、D118T+Y242F+A185Q、D118T+Y242F+N194F、D118T+E259G+F266Y、D118T+E259G+N279S、D118T+E259G+K280Y、D118T+E259G+G303R、D118T+E259G+E390A、D118T+E259G+G475K、D118T+E259G+A185Q、D118T+E259G+N194F、D118T+F266Y+N279S、D118T+F266Y+K280Y、D118T+F266Y+G303R、D118T+F266Y+E390A、D118T+F266Y+G475K、D118T+F266Y+A185Q、D118T+F266Y+N194F、D118T+N279S+K280Y、D118T+N279S+G303R、D118T+N279S+E390A、D118T+N279S+G475K、D118T+N279S+A185Q、D118T+N279S+N194F、D118T+K280Y+G303R、D118T+K280Y+E390A、D118T+K280Y+G475K、D118T+K280Y+A185Q、D118T+K280Y+N194F、D118T+G303R+E390A、D118T+G303R+G475K、D118T+G303R+A185Q、D118T+G303R+N194F、D118T+E390A+G475K、D118T+E390A+A185Q、D118T+E390A+N194F、D118T+G475K+A185Q、D118T+G475K+N194F、D118T+A185Q+N194F、W140Y+T182S+A185D、W140Y+T182S+E189P、W140Y+T182S+Y242F、W140Y+T182S+E259G、W140Y+T182S+F266Y、W140Y+T182S+N279S、W140Y+T182S+K280Y、W140Y+T182S+G303R、W140Y+T182S+E390A、W140Y+T182S+G475K、W140Y+T182S+A185Q、W140Y+T182S+N194F、W140Y+A185D+E189P、W140Y+A185D+Y242F、W140Y+A185D+E259G、W140Y+A185D+F266Y、W140Y+A185D+N279S、W140Y+A185D+K280Y、W140Y+A185D+G303R、W140Y+A185D+E390A、W140Y+A185D+G475K、W140Y+A185D+A185Q、W140Y+A185D+N194F、W140Y+E189P+Y242F、W140Y+E189P+E259G、W140Y+E189P+F266Y、W140Y+E189P+N279S、W140Y+E189P+K280Y、W140Y+E189P+G303R、W140Y+E189P+E390A、W140Y+E189P+G475K、W140Y+E189P+A185Q、W140Y+E189P+N194F、W140Y+Y242F+E259G、W140Y+Y242F+F266Y、W140Y+Y242F+N279S、W140Y+Y242F+K280Y、W140Y+Y242F+G303R、W140Y+Y242F+E390A、W140Y+Y242F+G475K、W140Y+Y242F+A185Q、W140Y+Y242F+N194F、W140Y+E259G+F266Y、W140Y+E259G+N279S、W140Y+E259G+K280Y、W140Y+E259G+G303R、W140Y+E259G+E390A、W140Y+E259G+G475K、W140Y+E259G+A185Q、W140Y+E259G+N194F、W140Y+F266Y+N279S、W140Y+F266Y+K280Y、W140Y+F266Y+G303R、W140Y+F266Y+E390A、W140Y+F266Y+G475K、W140Y+F266Y+A185Q、W140Y+F266Y+N194F、W140Y+N279S+K280Y、W140Y+N279S+G303R、W140Y+N279S+E390A、W140Y+N279S+G475K、W140Y+N279S+A185Q、W140Y+N279S+N194F、W140Y+K280Y+G303R、W140Y+K280Y+E390A、W140Y+K280Y+G475K、W140Y+K280Y+A185Q、W140Y+K280Y+N194F、W140Y+G303R+E390A、W140Y+G303R+G475K、W140Y+G303R+A185Q、W140Y+G303R+N194F、W140Y+E390A+G475K、W140Y+E390A+A185Q、W140Y+E390A+N194F、W140Y+G475K+A185Q、W140Y+G475K+N194F、W140Y+A185Q+N194F、T182S+A185D+E189P、T182S+A185D+Y242F、T182S+A185D+E259G、T182S+A185D+F266Y、T182S+A185D+N279S、T182S+A185D+K280Y、T182S+A185D+G303R、T182S+A185D+E390A、T182S+A185D+G45K、T182S+A185D+A185Q、T182S+A185D+N194F、T182S+E189P+Y242F、T182S+E189P+E259G、T182S+E189P+F266Y、T182S+E189P+N279S、T182S+E189P+K280Y、T182S+E189P+G303R、T182S+E189P+E390A、T182S+E189P+G475K、T182S+E189P+A185Q、T182S+E189P+N194F、T1、 82S+Y242F+E259G、T182S+Y242F+F266Y、T182S+Y242F+N279S、T182S+Y242F+K280Y、T182S+Y242F+G303R、T182S+Y242F+E390A、T182S+Y242F+G475K、T182S+Y242F+A185Q、T182S+Y242F+N194F、T182S+E259G+F266Y、T182S+E259G+N279S、T182S+E259G+K280Y、T182S+E259G+G303R、T182S+E259G+E390A、T182S+E259G+G475K、T182S+E259G+A185Q、T182S+E259G+N194F、T182S+F266Y+N279S、T182S+F266Y+K280Y、T182S+F266Y+G303R、T182S+F266Y+E390A、T182S+F266Y+G475K、T182S+F266Y+A185Q、T182S+F266Y+N194F、T182S+N279S+K280Y、T182S+N279S+G303R、T182S+N279S+E390A、T182S+N279S+G475K、T182S+N279S+A185Q、T182S+N279S+N194F、T182S+K280Y+G303R、T182S+K280Y+E390A、T182S+K280Y+G475K、T182S+K280Y+A185Q、T182S+K280Y+N194F、T182S+G303R+E390A、T182S+G303R+G475K、T182S+G303R+A185Q、T182S+G303R+N194F、T182S+E390A+G475K、T182S+E390A+A185Q、T182S+E390A+N194F、T182S+G475K+A185Q、T182S+G475K+N194F、T182S+A185Q+N194F、A185D+E189P+Y242F、A185D+E189P+E259G、A185D+E189P+F266Y、A185D+E189P+N279S、A185D+E189P+K280Y、A185D+E189P+G303R、A185D+E189P+E390A、A185D+E189P+G475K、A185D+E189P+A185Q、A185D+E189P+N194F、A185D+Y242F+E259G、A185D+Y242F+F266Y、A185D+Y242F+N279S、A185D+Y242F+K280Y、A185D+Y242F+G303R、A185D+Y242F+E390A、A185D+Y242F+G475K、A185D+Y242F+A185Q、A185D+Y242F+N194F、A185D+E259G+F266Y、A185D+E259G+N279S、A185D+E259G+K280Y、A185D+E259G+G303R、A185D+E259G+E390A、A185D+E259G+G475K、A185D+E259G+A185Q、A185D+E259G+N194F、A185D+F266Y+N279S、A185D+F266Y+K280Y、A185D+F266Y+G303R、A185D+F266Y+E390A、A185D+F266Y+G475K、A185D+F266Y+A185Q、A185D+F266Y+N194F、A185D+N279S+K280Y、A185D+N279S+G303R、A185D+N279S+E390A、A185D+N279S+G475K、A185D+N279S+A185Q、A185D+N279S+N194F、A185D+K280Y+G303R、A185D+K280Y+E390A、A185D+K280Y+G475K、A185D+K280Y+A185Q、A185D+K280Y+N194F、A185D+G303R+E390A、A185D+G303R+G475K、A185D+G303R+A185Q、A185D+G303R+N194F、A185D+E390A+G475K、A185D+E390A+A185Q、A185D+E390A+N194F、A185D+G475K+A185Q、A185D+G475K+N194F、A185D+A185Q+N194F、E189P+Y242F+E259G、E189P+Y242F+F266Y、E189P+Y242F+N279S、E189P+Y242F+K280Y、E189P+Y242F+G303R、E189P+Y242F+E390A、E189P+Y242F+G475K、E189P+Y242F+A185Q、E189P+Y242F+N194F、E189P+E259G+F266Y、E189P+E259G+N279S、E189P+E259G+K280Y、E189P+E259G+G303R、E189P+E259G+E390A、E189P+E259G+G475K、E189P+E259G+A185Q、E189P+E259G+N194F、E189P+F266Y+N279S、E189P+F266Y+K280Y、E189P+F266Y+G303R、E189P+F266Y+E390A、E189P+F266Y+G475K、E189P+F266Y+A185Q、E189P+F266Y+N194F、E189P+N279S+K280Y、E189P+N279S+G303R、E189P+N279S+E390A、E189P+N279S+G475K、E189P+N279S+A185Q、E189P+N279S+N194F、E189P+K280Y+G303R、E189P+K280Y+E390A、E189P+K280Y+G475K、E189P+K280Y+A185Q、E189P+K280Y+N194F、E189P+G303R+E390A、E189P+G303R+G475K、E189P+G303R+A185Q、E189P+G303R+N194F、E189P+E390A+G475K、E189P+E390A+A185Q、E189P+E390A+N194F、E189P+G475K+A185Q、E189P+G475K+N194F、E189P+A185Q+N194F、Y242F+E259G+F266Y、Y242F+E259G+N279S、Y242F+E259G+K280Y、Y242F+E259G+G303R、Y242F+E259G+E390A、Y242F+E259G+G475K、Y242F+E259G+A185Q、Y242F+E259G+N194F、Y242F+F266Y+N279S、Y242F+F266Y+K280Y、Y242F+F266Y+G303R、Y242F+F266Y+E390A、Y242F+F266Y+G475K、Y242F+F266Y+A185Q、Y242F+F266Y+N194F、Y242F+N279S+K280Y、Y242F+N279S+G303R、Y242F+N279S+E390A、Y242F+N279S+G475K、Y242F+N279S+A185Q、Y242F+N279S+N194F、Y242F+K280Y+G303R、Y242F+K280Y+E390A、Y242F+K 280Y+G475K、Y242F+K280Y+A185Q、Y242F+K280Y+N194F、Y242F+G303R+E39 0A、Y242F+G303R+G475K、Y242F+G303R+A185Q、Y242F+G303R+N194F、Y242F +E390A+G475K、Y242F+E390A+A185Q、Y242F+E390A+N194F、Y242F+G475K+A 185Q、Y242F+G475K+N194F、Y242F+A185Q+N194F、E259G+F266Y+N279S、E25 9G+F266Y+K280Y、E259G+F266Y+G303R、E259G+F266Y+E390A、E259G+F266Y +G475K, E259G+F266Y+A185Q, E259G+F266Y+N194F, E259G+N279S+K280Y, E259G+N279S+G303R, E259G+N279S+E390A, E259G+N279S+G475K, E259G+N27 9S+A185Q, E259G+N279S+N194F, E259G+K280Y+G303R, E259G+K280Y+E390A, E259G+K280Y+G475K, E259G+K280Y+A185Q, E259G+K280Y+N194F, E259G+ G303R+E390A, E259G+G303R+G475K, E259G+G303R+A185Q, E259G+G303R+N194F, E259G+E390A+G475K, E259G+E390A+A185Q, E259G+E390A+N194F, E259 G+G475K+A185Q, E259G+G475K+N194F, E259G+A185Q+N194F, F266Y+N279S+K280Y, F266Y+N279S+G303R, F266Y+N279S+E390A, F266Y+N279S+G475K, F2 66Y+N279S+A185Q、F266Y+N279S+N194F、F266Y+K280Y+G303R、F266Y+K280 Y+E390A、F266Y+K280Y+G475K、F266Y+K280Y+A185Q、F266Y+K280Y+N194F、F266Y+G303R+E390A、F266Y+G303R+G475K、F266Y+G303R+A185Q、F266Y+G303R+N194F、F266Y+E390A+G475K、F266Y+E390A+A185Q、F266Y+E390A+N194F、F266Y+G475K+A185Q、F266Y+G475K+N194F、F266Y+A185Q+N194F、N279S+K280Y+G303R、N279S+K280Y+E390A、N279S+K280Y+G475K、N279S+K280Y+A185Q、N279S+K280Y+N194F、N279S+G303R+E390A、N279S+G303R+G475K、N279S+G303R+A185Q、N279S+G303R+N194F、N279S+E390A+G475K、N279S+E390A+A185Q、N279S+E390A+N194F、N279S+G475K+A185Q、 N279S+G475K+N194F, N279S+A185Q+N194F, K280Y+G303R+E390A, K280Y+G303R+G475K, K280Y+G303R+A185Q , K280Y+G303R+N194F, K280Y+E390A+G475K, K280Y+E390A+A185Q, K280Y+E390A+N194F, K280Y+G475K+A185 Q, K280Y+G475K+N194F, K280Y+A185Q+N194F, G303R+E390A+G475K, G303R+E390A+A185Q, G303R+E390A+N19 4F, G303R+G475K+A185Q, G303R+G475K+N194F, G303R+A185Q+N194F, E390A+G475K+A185Q, E390A+G475K+N1 The variant includes modifications at positions corresponding to 94F, E390A+A185Q+N194F, and G475K+A185Q+N194F, wherein the variant has sequence homology with SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 or 14 of at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%, and the variant has α-amylase activity.

[0377] In one embodiment, the present invention relates to an α-amylase variant of SEQ ID NO: 1, a) Using sequence number 1 for numbering, deletes are made at positions corresponding to R180* and S181*, b) Use array index 1 for numbering: H1*+G7A+W140Y+E189P, H1*+G7A+W140Y+Y242F, H1*+G7A+W140Y+E259G, H1*+G7A+W140Y+N279S, H1*+G7A+W140Y+K280Y, H1*+G7A+W140Y+G303R, H1*+G7A+W140Y+E390A, H1*+G7A+E189P+Y242F, H1*+G7A+E189P+E259G, H1*+G7A+E189P+N279S, H1*+G7A+E189P+K280Y, H1*+G7A+E1 89P+G303R, H1*+G7A+E189P+E390A, H1*+G7A+Y242F+E259G, H1*+G7A+Y242 F+N279S, H1*+G7A+Y242F+K280Y, H1*+G7A+Y242F+G303R, H1*+G7A+Y242F+ E390A, H1*+G7A+E259G+N279S, H1*+G7A+E259G+K280Y, H1*+G7A+E259G+G3 03R, H1*+G7A+E259G+E390A, H1*+G7A+N279S+K280Y, H1*+G7A+N279S+G303R , H1*+G7A+N279S+E390A, H1*+G7A+K280Y+G303R, H1*+G7A+K280Y+E390A, H 1*+G7A+G303R+E390A, H1*+W140Y+E189P+Y242F, H1*+W140Y+E189P+E259G , H1*+W140Y+E189P+N279S, H1*+W140Y+E189P+K280Y, H1*+W140Y+E189P+G 303R, H1*+W140Y+E189P+E390A, H1*+W140Y+Y242F+E259G, H1*+W140Y+Y242 F+N279S, H1*+W140Y+Y242F+K280Y, H1*+W140Y+Y242F+G303R, H1*+W140Y+ Y242F+E390A, H1*+W140Y+E259G+N279S, H1*+W140Y+E259G+K280Y, H1*+W14 0Y+E259G+G303R, H1*+W140Y+E259G+E390A, H1*+W140Y+N279S+K280Y, H1* +W140Y+N279S+G303R, H1*+W140Y+N279S+E390A, H1*+W140Y+K280Y+G303R,H1*+W140Y+K280Y+E390A、H1*+W140Y+G303R+E390A、H1*+E189P+Y242F+E259G、H1*+E189P+Y242F+N279S、H1*+E189P+Y242F+K280Y、H1*+E189P+Y242F+G303R、H1*+E189P+Y242F+E390A、H1*+E189P+E259G+N279S、H1*+E189P+E259G+K280Y、H1*+E189P+E259G+G303R、H1*+E189P+E259G+E390A、H1*+E189P+N279S+K280Y、H1*+E189P+N279S+G303R、H1*+E189P+N279S+E390A、H1*+E189P+K280Y+G303R、H1*+E189P+K280Y+E390A、H1*+E189P+G303R+E390A、H1*+Y242F+E259G+N279S、H1*+Y242F+E259G+K280Y、H1*+Y242F+E259G+G303R、H1*+Y242F+E259G+E390A、H1*+Y242F+N279S+K280Y、H1*+Y242F+N279S+G303R、H1*+Y242F+N279S+E390A、H1*+Y242F+K280Y+G303R、H1*+Y242F+K280Y+E390A、H1*+Y242F+G303R+E390A、H1*+E259G+N279S+K280Y、H1*+E259G+N279S+G303R、H1*+E259G+N279S+E390A、H1*+E259G+K280Y+G303R、H1*+E259G+K280Y+E390A、H1*+E259G+G303R+E390A、H1*+N279S+K280Y+G303R、H1*+N279S+K280Y+E390A、H1*+N279S+G303R+E390A、H1*+K280Y+G303R+E390A、G7A+W140Y+E189P+Y242F、G7A+W140Y+E189P+E259G、G7A+W140Y+E189P+N279S、G7A+W140Y+E189P+K280Y、G7A+W140Y+E189P+G303R、G7A+W140Y+E189P+E390A、G7A+W140Y+Y242F+E259G、G7A+W140Y+Y242F+N279S、G7A+W140Y+Y242F+K280Y, G7A+W140Y+Y242F+G303R, G7A+W140Y+Y242F+E390A, G7A+W140Y+E259G+N279S, G7A+W140Y+E259G+K280Y, G7A+W140Y+E25 9G+G303R、G7A+W140Y+E259G+E390A、G7A+W140Y+N279S+K280Y、G7A+W140Y +N279S+G303R、G7A+W140Y+N279S+E390A、G7A+W140Y+K280Y+G303R、G7A+W 140Y+K280Y+E390A, G7A+W140Y+G303R+E390A, G7A+E189P+Y242F+E259G, G7A+E189P+Y242F+N279S, G7A+E189P+Y242F+K280Y, G7A+E189P+Y242F+G30 3R, G7A+E189P+Y242F+E390A, G7A+E189P+E259G+N279S, G7A+E189P+E259G+K280Y, G7A+E189P+E259G+G303R, G7A+E189P+E259G+E390A, G7A+E189P+N 279S+K280Y、G7A+E189P+N279S+G303R、G7A+E189P+N279S+E390A、G7A+E1 89P+K280Y+G303R、G7A+E189P+K280Y+E390A、G7A+E189P+G303R+E390A、G7 A+Y242F+E259G+N279S, G7A+Y242F+E259G+K280Y, G7A+Y242F+E259G+G303R, G7A+Y242F+E259G+E390A, G7A+Y242F+N279S+K280Y, G7A+Y242F+N279S+ G303R, G7A+Y242F+N279S+E390A, G7A+Y242F+K280Y+G303R, G7A+Y242F+K280Y+E390A, G7A+Y242F+G303R+E390A, G7A+E259G+N279S+K280Y, G7A+E259 G+N279S+G303R, G7A+E259G+N279S+E390A, G7A+E259G+K280Y+G303R, G7A+E259G+K280Y+E390A, G7A+E259G+G303R+E390A, G7A+N279S+K280Y+G303RG7A+N279S+K280Y+E390A、G7A+N279S+G303R+E390A、G7A+K280Y+G303R+E390A、W140Y+E189P+Y242F+E259G、W140Y+E189P+Y242F+N279S、W140Y+E189P+Y242F+K280Y、W140Y+E189P+Y242F+G303R、W140Y+E189P+Y242F+E390A、W140Y+E189P+E259G+N279S、W140Y+E189P+E259G+K280Y、W140Y+E189P+E259G+G303R、W140Y+E189P+E259G+E390A、W140Y+E189P+N279S+K280Y、W140Y+E189P+N279S+G303R、W140Y+E189P+N279S+E390A、W140Y+E189P+K280Y+G303R、W140Y+E189P+K280Y+E390A、W140Y+E189P+G303R+E390A、W140Y+Y242F+E259G+N279S、W140Y+Y242F+E259G+K280Y、W140Y+Y242F+E259G+G303R、W140Y+Y242F+E259G+E390A、W140Y+Y242F+N279S+K280Y、W140Y+Y242F+N279S+G303R、W140Y+Y242F+N279S+E390A、W140Y+Y242F+K280Y+G303R、W140Y+Y242F+K280Y+E390A、W140Y+Y242F+G303R+E390A、W140Y+E259G+N279S+K280Y、W140Y+E259G+N279S+G303R、W140Y+E259G+N279S+E390A、W140Y+E259G+K280Y+G303R、W140Y+E259G+K280Y+E390A、W140Y+E259G+G303R+E390A、W140Y+N279S+K280Y+G303R、W140Y+N279S+K280Y+E390AW140Y+N279S+G303R+E390A、W140Y+K280Y+G303R+E390A、E189P+Y242F+E259G+N279S、E189P+Y242F+E259G+K280Y、E189P+Y242F+E259G+G303R、E189P+Y242F+E259G+E390A、E189P+Y242F+N279S+K280Y, E189P+Y242F+N279S+G303R, E189P+Y242F+N2 79S+E390A, E189P+Y242F+K280Y+G303R, E189P+Y242F+K280Y+E390A, E189 P+Y242F+G303R+E390A, E189P+E259G+N279S+K280Y, E189P+E259G+N279S+ G303R, E189P+E259G+N279S+E390A, E189P+E259G+K280Y+G303R, E189P+E2 59G+K280Y+E390A, E189P+E259G+G303R+E390A, E189P+N279S+K280Y+G303 R, E189P+N279S+K280Y+E390A, E189P+N279S+G303R+E390A, E189P+K280Y+ G303R+E390A, Y242F+E259G+N279S+K280Y, Y242F+E259G+N279S+G303R, Y2 42F+E259G+N279S+E390A, Y242F+E259G+K280Y+G303R, Y242F+E259G+K280Y +E390A, Y242F+E259G+G303R+E390A, Y242F+N279S+K280Y+G303R, Y242F+N 279S+K280Y+E390A, Y242F+N279S+G303R+E390A, Y242F+K280Y+G303R+E39 0A, E259G+N279S+K280Y+G303R, E259G+N279S+K280Y+E390A, E259G+N279S+G303R+E390A, E259G+K280Y+G303R+E390A, N279S+K280Y+G303R+E390A The variant includes modifications in the corresponding positions, and the variant is sequence numbers 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 or 14, and at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%,The variants possess at least 97%, at least 98%, or at least 99%, but less than 100%, sequence homology, and the variants exhibit α-amylase activity.

[0378] In one embodiment, the α-amylase mutant of SEQ ID NO: 1 is a) Using sequence number 1 for numbering, deletes are made at positions corresponding to R180* and S181*, b) Using sequence number 1 for numbering: H1*+G7A+W140Y+E189P+Y242F, H1*+G7A+W140Y+E189P+E259G, H1*+G7A+W140Y+E189P+N279S, H1*+G7A+W140Y+E189P+K280Y, H1*+G7A+W140Y+E189P+G303R, H1*+G7A+W140Y+E189P+E390A, H1*+G7A+W140Y+Y242F+E259G, H1*+G7A+W140Y+Y242F+N279S, H1*+G7A+W140Y+Y242 F+K280Y, H1*+G7A+W140Y+Y242F+G303R, H1*+G7A+W140Y+Y242F+E390A, H 1*+G7A+W140Y+E259G+N279S, H1*+G7A+W140Y+E259G+K280YH1*+G7A+W14 0Y+E259G+G303R, H1*+G7A+W140Y+E259G+E390A, H1*+G7A+W140Y+N279S+ K280Y, H1*+G7A+W140Y+N279S+G303R, H1*+G7A+W140Y+N279S+E390A, H1*+ G7A+W140Y+K280Y+G303R, H1*+G7A+W140Y+K280Y+E390A, H1*+G7A+W140Y +G303R+E390A, H1*+G7A+E189P+Y242F+E259G, H1*+G7A+E189P+Y242F+N2 79S, H1*+G7A+E189P+Y242F+K280Y, H1*+G7A+E189P+Y242F+G303R, H1*+G 7A+E189P+Y242F+E390A, H1*+G7A+E189P+E259G+N279S, H1*+G7A+E189P+E 259G+K280Y, H1*+G7A+E189P+E259G+G303R, H1*+G7A+E189P+E259G+E390 A, H1*+G7A+E189P+N279S+K280Y, H1*+G7A+E189P+N279S+G303R, H1*+G7A +E189P+N279S+E390A, H1*+G7A+E189P+K280Y+G303R, H1*+G7A+E189P+K2 80Y+E390A, H1*+G7A+E189P+G303R+E390A, H1*+G7A+Y242F+E259G+N279S,H1*+G7A+Y242F+E259G+K280Y、H1*+G7A+Y242F+E259G+G303R、H1*+G7A+Y242F+E259G+E390A、H1*+G7A+Y242F+N279S+K280Y、H1*+G7A+Y242F+N279S+G303R、H1*+G7A+Y242F+N279S+E390A、H1*+G7A+Y242F+K280Y+G303R、H1*+G7A+Y242F+K280Y+E390A、H1*+G7A+Y242F+G303R+E390A、H1*+G7A+E259G+N279S+K280Y、H1*+G7A+E259G+N279S+G303R、H1*+G7A+E259G+N279S+E390A、H1*+G7A+E259G+K280Y+G303R、H1*+G7A+E259G+K280Y+E390A、H1*+G7A+E259G+G303R+E390A、H1*+G7A+N279S+K280Y+G303R、H1*+G7A+N279S+K280Y+E390A、H1*+G7A+N279S+G303R+E390A、H1*+G7A+K280Y+G303R+E390A、H1*+W140Y+E189P+Y242F+E259G、H1*+W140Y+E189P+Y242F+N279S、H1*+W140Y+E189P+Y242F+K280Y、H1*+W140Y+E189P+Y242F+G303R、H1*+W140Y+E189P+Y242F+E390A、H1*+W140Y+E189P+E259G+N279S、H1*+W140Y+E189P+E259G+K280Y、H1*+W140Y+E189P+E259G+G303R、H1*+W140Y+E189P+E259G+E390A、H1*+W140Y+E189P+N279S+K280Y、H1*+W140Y+E189P+N279S+G303R、H1*+W140Y+E189P+N279S+E390A、H1*+W140Y+E189P+K280Y+G303R、H1*+W140Y+E189P+K280Y+E390A、H1*+W140Y+E189P+G303R+E390A、H1*+W140Y+Y242F+E259G+N279S、H1*+W140Y+Y242F+E259G+K280Y、H1*+W140Y+Y242F+E259G+G303R、H1*+W140Y+Y242F+E259G+E390A、H1*+W140Y+Y242F+N279S+K280Y、H1*+W140Y+Y242F+N279S+G303R、H1*+W140Y+Y242F+N279S+E390A、H1*+W140Y+Y242F+K280Y+G303R、H1*+W140Y+Y242F+K280Y+E390A、H1*+W140Y+Y242F+G303R+E390A、H1*+W140Y+E259G+N279S+K280Y、H1*+W140Y+E259G+N279S+G303R、H1*+W140Y+E259G+N279S+E390A、H1*+W140Y+E259G+K280Y+G303R、H1*+W140Y+E259G+K280Y+E390A、H1*+W140Y+E259G+G303R+E390A、H1*+W140Y+N279S+K280Y+G303R、H1*+W140Y+N279S+K280Y+E390A、H1*+W140Y+N279S+G303R+E390A、H1*+W140Y+K280Y+G303R+E390A、H1*+E189P+Y242F+E259G+N279S、H1*+E189P+Y242F+E259G+K280Y、H1*+E189P+Y242F+E259G+G303R、H1*+E189P+Y242F+E259G+E390A、H1*+E189P+Y242F+N279S+K280Y、H1*+E189P+Y242F+N279S+G303R、H1*+E189P+Y242F+N279S+E390A、H1*+E189P+Y242F+K280Y+G303R、H1*+E189P+Y242F+K280Y+E390A、H1*+E189P+Y242F+G303R+E390A、H1*+E189P+E259G+N279S+K280Y、H1*+E189P+E259G+N279S+G303R、H1*+E189P+E259G+N279S+E390A、H1*+E189P+E259G+K280Y+G303R、H1*+E189P+E259G+K280Y+E390A、H1*+E189P+E259G+G303R+E390A、H1*+E189P+N279S+K280Y+G303R、H1*+E189P+N279S+K280Y+E390A、H1*+E189P+N279S+G303R+E390A、H1*+E189P+K280Y+G303R+E390A、H1*+Y242F+E259G+N279S+K280Y、H1*+Y242F+E259G+N279S+G303R、H1*+Y242F+ E259G+N279S+E390A, H1*+Y242F+E259G+K280Y+G303R, H1*+Y242F+E259G+K280Y+E390A, H1*+Y242F+E259G+G303R+E390A, H1*+Y242F+N279S+K280Y +G303R, H1*+Y242F+N279S+K280Y+E390A, H1*+Y242F+N279S+G303R+E390A, H1*+Y242F+K280Y+G303R+E390A, H1*+E259G+N279S+K280Y+G303R, H1* +E259G+N279S+K280Y+E390A、H1*+E259G+N279S+G303R+E390A、H1*+E259G+K280Y+G303R+E390A、H1*+N279S+K280Y+G303R+E390A、G7A+W140Y+E189 P+Y242F+E259G, G7A+W140Y+E189P+Y242F+N279S, G7A+W140Y+E189P+Y242F+K280Y, G7A+W140Y+E189P+Y242F+G303R, G7A+W140Y+E189P+Y242F+E3 90A、G7A+W140Y+E189P+E259G+N279S、G7A+W140Y+E189P+E259G+K280Y、G 7A+W140Y+E189P+E259G+G303R、G7A+W140Y+E189P+E259G+E390A、G7A+W14 0Y+E189P+N279S+K280Y、G7A+W140Y+E189P+N279S+G303R、G7A+W140Y+E1 89P+N279S+E390A、G7A+W140Y+E189P+K280Y+G303R、G7A+W140Y+E189P+K 280Y+E390A, G7A+W140Y+E189P+G303R+E390A, G7A+W140Y+Y242F+E259G+N279S, G7A+W140Y+Y242F+E259G+K280Y, G7A+W140Y+Y242F+E259G+G303RG7A+W140Y+Y242F+E259G+E390A, G7A+W140Y+Y242F+N279S+K280Y, G7A+W140Y+Y242F+N279S+G303R, G7A+W140Y+Y242F+N279S+E390A, G7A+W140Y+ Y242F+K280Y+G303R, G7A+W140Y+Y242F+K280Y+E390A, G7A+W140Y+Y242F+G303R+E390A, G7A+W140Y+E259G+N279S+K280Y, G7A+W140Y+E259G+N279S +G303R, G7A+W140Y+E259G+N279S+E390A, G7A+W140Y+E259G+K280Y+G303R, G7A+W140Y+E259G+K280Y+E390A, G7A+W140Y+E259G+G303R+E390A, G7A +W140Y+N279S+K280Y+G303R、G7A+W140Y+N279S+K280Y+E390A、G7A+W140Y+N279S+G303R+E390A、G7A+W140Y+K280Y+G303R+E390A、G7A+E189P+Y242 F+E259G+N279S、G7A+E189P+Y242F+E259G+K280Y、G7A+E189P+Y242F+E25 9G+G303R、G7A+E189P+Y242F+E259G+E390A、G7A+E189P+Y242F+N279S+K2 80Y, G7A+E189P+Y242F+N279S+G303R, G7A+E189P+Y242F+N279S+E390A, G7A+E189P+Y242F+K280Y+G303R, G7A+E189P+Y242F+K280Y+E390A, G7A+E18 9P+Y242F+G303R+E390A、G7A+E189P+E259G+N279S+K280Y、G7A+E189P+E2 59G+N279S+G303R、G7A+E189P+E259G+N279S+E390A、G7A+E189P+E259G+K 280Y+G303R, G7A+E189P+E259G+K280Y+E390A, G7A+E189P+E259G+G303R+E390A, G7A+E189P+N279S+K280Y+G303R, G7A+E189P+N279S+K280Y+E390AG7A+E189P+N279S+G303R+E390A, G7A+E189P+K280Y+G303R+E390A, G7A+Y242F+E259G+N279S+K280Y, G7A+Y242F+E259G+N279S+G303R, G7A+Y242F+E259G+N279S+E390A, G7A+Y242F+E259G+K280Y+G303R, G7A+Y242F+E259G+K280Y+E390A, G7A+Y2 42F+E259G+G303R+E390A、G7A+Y242F+N279S+K280Y+G303R、G7A+Y242F+N279S+K280Y+E390A、G7A+Y242F+N279S+G303R+E390A、G7A+Y242F+K280Y+G303R+E390A、G7A+E259G+N279S+K280Y+G303R、G7A+E259G+N279S+K280Y+E390A、G7A+E259G+N279S+G303R+E390A、G7A+E259G+K280Y+G303R+E390A、G7A+N279S+K280Y+G303R+E390A、W140Y+E189P+Y242F+E259G+N279S、W140Y+E189P+Y242F+E259G+K280Y、W140Y+E189P+Y242F+E259G+G303R、W140Y+E189P+Y242F+E259G+E390A、W140Y+E189P+Y242F+N279S+K280Y、W140Y+E189P+Y242F+N279S+G303R、W140Y+E189P+Y242F+N279S+E390A、W140Y+E189P+Y242F+K280Y+G303R、W140Y+E189P+Y242F+K280Y+E390A、W140Y+E189P+Y242F+G303R+E390A、W140Y+E189P+E259G+N279S+K280Y、W140Y+E189P+E259G+N279S+G303R、W140Y+E189P+E259G+N279S+E390A、W140Y+E189P+E259G+K280Y+G303R、W140Y+E189P+E259G+K280Y+E390A、W140Y+E189P+E259G+G303R+E390A、W140Y+E189P+N279S+K280Y+G303R、W140Y+E189P+N279S+K280Y+E390A、W140Y+E189P+N279S+G303R+E390A、W140Y+E189P+K280Y+G303R+E390A、W140Y+Y242F+E259G+N279S+K280Y、W140Y+Y242F+E259G+N279S+G303R、W140Y+Y242F+E259G+N279S+E390A、W140Y+Y242F+E259G+K280Y+G303R、W140Y+Y242F+E259G+K280Y+E390A, W140Y+Y242F+E259G+G303R+E390A, W1 40Y+Y242F+N279S+K280Y+G303R, W140Y+Y242F+N279S+K280Y+E390A, W140Y +Y242F+N279S+G303R+E390A, W140Y+Y242F+K280Y+G303R+E390A, W140Y+E 259G+N279S+K280Y+G303R, W140Y+E259G+N279S+K280Y+E390A, W140Y+E259 G+N279S+G303R+E390A, W140Y+E259G+K280Y+G303R+E390A, W140Y+N279S+ K280Y+G303R+E390A, E189P+Y242F+E259G+N279S+K280Y, E189P+Y242F+E25 9G+N279S+G303R, E189P+Y242F+E259G+N279S+E390A, E189P+Y242F+E259G +K280Y+G303R, E189P+Y242F+E259G+K280Y+E390A, E189P+Y242F+E259G+G3 03R+E390A, E189P+Y242F+N279S+K280Y+G303R, E189P+Y242F+N279S+K280 Y+E390A, E189P+Y242F+N279S+G303R+E390A, E189P+Y242F+K280Y+G303R+E 390A, E189P+E259G+N279S+K280Y+G303R, E189P+E259G+N279S+K280Y+E39 0A, E189P+E259G+N279S+G303R+E390A, E189P+E259G+K280Y+G303R+E390A, The variants include modifications at positions corresponding to E189P+N279S+K280Y+G303R+E390A, Y242F+E259G+N279S+K280Y+G303R, Y242F+E259G+N279S+K280Y+E390A, Y242F+E259G+N279S+G303R+E390A, Y242F+E259G+K280Y+G303R+E390A, and the variants include modifications at positions corresponding to sequence numbers 1, 2, 3, 4, 5, 6, 7,The variant has at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%, and the variant has α-amylase activity.

[0379] In one embodiment, the α-amylase mutant of SEQ ID NO: 1 is a) Using sequence number 1 for numbering, deletes are made at positions corresponding to R180* and S181*, b) Using sequence number 1 for numbering: H1*+G7A+W140Y+E189P+Y242F+E259G, H1*+G7A+W140Y+E189P+Y242F+N279S, H1*+G7A+W140Y+E189P+Y242F+K280Y, H1*+G7A+W140Y+E189P+Y242F+G303R, H1*+G7A+W140Y+E189P+Y242F+E390A, H1*+G7A+W140Y+E189P+E259G+N279S, H1*+G7A+W140Y+E189P+E259G+K280Y, H 1*+G7A+W140Y+E189P+E259G+G303R, H1*+G7A+W140Y+E189P+E259G+E390 A, H1*+G7A+W140Y+E189P+N279S+K280Y, H1*+G7A+W140Y+E189P+N279S+G 303R, H1*+G7A+W140Y+E189P+N279S+E390A, H1*+G7A+W140Y+E189P+K280 Y+G303R, H1*+G7A+W140Y+E189P+K280Y+E390A, H1*+G7A+W140Y+E189P+G3 03R+E390A, H1*+G7A+W140Y+Y242F+E259G+N279S, H1*+G7A+W140Y+Y242F +E259G+K280Y, H1*+G7A+W140Y+Y242F+E259G+G303R, H1*+G7A+W140Y+Y2 42F+E259G+E390A, H1*+G7A+W140Y+Y242F+N279S+K280Y, H1*+G7A+W140Y +Y242F+N279S+G303R, H1*+G7A+W140Y+Y242F+N279S+E390A, H1*+G7A+W14 0Y+Y242F+K280Y+G303R, H1*+G7A+W140Y+Y242F+K280Y+E390A, H1*+G7A+ W140Y+Y242F+G303R+E390A, H1*+G7A+W140Y+E259G+N279S+K280Y, H1*+G 7A+W140Y+E259G+N279S+G303R, H1*+G7A+W140Y+E259G+N279S+E390A, H1 *+G7A+W140Y+E259G+K280Y+G303R, H1*+G7A+W140Y+E259G+K280Y+E390A,H1*+G7A+W140Y+E259G+G303R+E390A, H1*+G7A+W140Y+N279S+K280Y+G303R, H1*+G7A+W140Y+N279S+K280Y+E390A, H1*+G7A+W140Y+N279S+G303R+E3 90A, H1*+G7A+W140Y+K280Y+G303R+E390A, H1*+G7A+E189P+Y242F+E259G+N279S, H1*+G7A+E189P+Y242F+E259G+K280Y, H1*+G7A+E189P+Y242F+E259 G+G303R、H1*+G7A+E189P+Y242F+E259G+E390A、H1*+G7A+E189P+Y242F+N2 79S+K280Y、H1*+G7A+E189P+Y242F+N279S+G303R、H1*+G7A+E189P+Y242F+ N279S+E390A, H1*+G7A+E189P+Y242F+K280Y+G303R, H1*+G7A+E189P+Y242F+K280Y+E390A, H1*+G7A+E189P+Y242F+G303R+E390A, H1*+G7A+E189P+E2 59G+N279S+K280Y、H1*+G7A+E189P+E259G+N279S+G303R、H1*+G7A+E189P+E259G+N279S+E390A、H1*+G7A+E189P+E259G+K280Y+G303R、H1*+G7A+E189 P+E259G+K280Y+E390A、H1*+G7A+E189P+E259G+G303R+E390A、H1*+G7A+E1 89P+N279S+K280Y+G303R、H1*+G7A+E189P+N279S+K280Y+E390A、H1*+G7A+ E189P+N279S+G303R+E390A、H1*+G7A+E189P+K280Y+G303R+E390A、H1*+G7A+Y242F+E259G+N279S+K280Y、H1*+G7A+Y242F+E259G+N279S+G303R、H1*+ G7A+Y242F+E259G+N279S+E390A, H1*+G7A+Y242F+E259G+K280Y+G303R, H1*+G7A+Y242F+E259G+K280Y+E390A, H1*+G7A+Y242F+E259G+G303R+E390AH1*+G7A+Y242F+N279S+K280Y+G303R、H1*+G7A+Y242F+N279S+K280Y+E390A、H1*+G7A+Y242F+N279S+G303R+E390A、H1*+G7A+Y242F+K280Y+G303R+E390A、H1*+G7A+E259G+N279S+K280Y+G303R、H1*+G7A+E259G+N279S+K280Y+E390A、H1*+G7A+E259G+N279S+G303R+E390A、H1*+G7A+E259G+K280Y+G303R+E390A、H1*+G7A+N279S+K280Y+G303R+E390A、H1*+W140Y+E189P+Y242F+E259G+N279S、H1*+W140Y+E189P+Y242F+E259G+K280Y、H1*+W140Y+E189P+Y242F+E259G+G303R、H1*+W140Y+E189P+Y242F+E259G+E390A、H1*+W140Y+E189P+Y242F+N279S+K280Y、H1*+W140Y+E189P+Y242F+N279S+G303R、H1*+W140Y+E189P+Y242F+N279S+E390A、H1*+W140Y+E189P+Y242F+K280Y+G303R、H1*+W140Y+E189P+Y242F+K280Y+E390A、H1*+W140Y+E189P+Y242F+G303R+E390A、H1*+W140Y+E189P+E259G+N279S+K280Y、H1*+W140Y+E189P+E259G+N279S+G303R、H1*+W140Y+E189P+E259G+N279S+E390A、H1*+W140Y+E189P+E259G+K280Y+G303R、H1*+W140Y+E189P+E259G+K280Y+E390A、H1*+W140Y+E189P+E259G+G303R+E390A、H1*+W140Y+E189P+N279S+K280Y+G303R、H1*+W140Y+E189P+N279S+K280Y+E390A、H1*+W140Y+E189P+N279S+G303R+E390A、H1*+W140Y+E189P+K280Y+G303R+E390A、H1*+W140Y+Y242F+E259G+N279S+K280Y、H1*+W140Y+Y242F+E259G+N279S+G303R、H1*+W140Y+Y242F+E259G+N279S+E390A、H1*+W140Y+Y242F+E259G+K280Y+G303R、H1*+W140Y+Y242F+E259G+K280Y+E390A、H1*+W140Y+Y242F+E259G+G303R+E390A、H1*+W140Y+Y242F+N279S+K280Y+G303R、H1*+W140Y+Y242F+N279S+K280Y+E390A、H1*+W140Y+Y242F+N279S+G303R+E390A、H1*+W140Y+Y242F+K280Y+G303R+E390A、H1*+W140Y+E259G+N279S+K280Y+G303R、H1*+W140Y+E259G+N279S+K280Y+E390A、H1*+W140Y+E259G+N279S+G303R+E390A、H1*+W140Y+E259G+K280Y+G303R+E390A、H1*+W140Y+N279S+K280Y+G303R+E390A、H1*+E189P+Y242F+E259G+N279S+K280Y、H1*+E189P+Y242F+E259G+N279S+G303R、H1*+E189P+Y242F+E259G+N279S+E390A、H1*+E189P+Y242F+E259G+K280Y+G303R、H1*+E189P+Y242F+E259G+K280Y+E390A、H1*+E189P+Y242F+E259G+G303R+E390A、H1*+E189P+Y242F+N279S+K280Y+G303R、H1*+E189P+Y242F+N279S+K280Y+E390A、H1*+E189P+Y242F+N279S+G303R+E390A、H1*+E189P+Y242F+K280Y+G303R+E390A、H1*+E189P+E259G+N279S+K280Y+G303R、H1*+E189P+E259G+N279S+K280Y+E390A、H1*+E189P+E259G+N279S+G303R+E390A、H1*+E189P+E259G+K280Y+G303R+E390A、H1*+E189P+N279S+K280Y+G303R+E390A、H1*+Y242F+E259G+N279S+K280Y+G303R、H1*+Y242F+E259G+N279S+K280Y+E390A、H1*+Y242F+E259G+N279S +G303R+E390A、H1*+Y242F+E259G+K280Y+G303R+E390A、H1*+Y242F+N279S+K280Y+G303R+E390A、H1*+E259G+N279S+K280Y+G303R+E390A、G7A+W140 Y+E189P+Y242F+E259G+N279S, G7A+W140Y+E189P+Y242F+E259G+K280Y, G7A+W140Y+E189P+Y242F+E259G+G303R, G7A+W140Y+E189P+Y242F+E259G+E 390A, G7A+W140Y+E189P+Y242F+N279S+K280Y, G7A+W140Y+E189P+Y242F+N279S+G303R, G7A+W140Y+E189P+Y242F+N279S+E390A, G7A+W140Y+E189P+Y 242F+K280Y+G303R, G7A+W140Y+E189P+Y242F+K280Y+E390A, G7A+W140Y+E189P+Y242F+G303R+E390A, G7A+W140Y+E189P+E259G+N279S+K280Y, G7A+ W140Y+E189P+E259G+N279S+G303R、G7A+W140Y+E189P+E259G+N279S+E39 0A、G7A+W140Y+E189P+E259G+K280Y+G303R、G7A+W140Y+E189P+E259G+K28 0Y+E390A、G7A+W140Y+E189P+E259G+G303R+E390A、G7A+W140Y+E189P+N2 79S+K280Y+G303R、G7A+W140Y+E189P+N279S+K280Y+E390A、G7A+W140Y+E1 89P+N279S+G303R+E390A、G7A+W140Y+E189P+K280Y+G303R+E390A、G7A+W140Y+Y242F+E259G+N279S+K280Y、G7A+W140Y+Y242F+E259G+N279S+G303R、G7A+W140Y+Y242F+E259G+N279S+E390A、G7A+W140Y+Y242F+E259G+K280Y+G303R、G7A+W140Y+Y242F+E259G+K280Y+E390A、G7A+W140Y+Y242F+E259G+G303R+E390A、G7A+W140Y+Y242F+N279S+K280Y+G303R、G7A+W140Y+Y242F+N279S+K280Y+E390A、G7A+W、 140Y+Y242F+N279S+G303R+E390A, G7A+W140Y+Y242F+K280Y+G303R+E390A, G7A+W140Y+E259G+N279S+K280Y+G303R, G7A+W140Y+E259G+N279S+K28 0Y+E390A、G7A+W140Y+E259G+N279S+G303R+E390A、G7A+W140Y+E259G+K280Y+G303R+E390A、G7A+W140Y+N279S+K280Y+G303R+E390A、G7A+E189P+Y2 42F+E259G+N279S+K280Y, G7A+E189P+Y242F+E259G+N279S+G303R, G7A+E189P+Y242F+E259G+N279S+E390A, G7A+E189P+Y242F+E259G+K280Y+G303R G7A+E189P+Y242F+E259G+K280Y+E390A, G7A+E189P+Y242F+E259G+G303R+E390A, G7A+E189P+Y242F+N279S+K280Y+G303R, G7A+E189P+Y242F+N279 S+K280Y+E390A、G7A+E189P+Y242F+N279S+G303R+E390A、G7A+E189P+Y24 2F+K280Y+G303R+E390A、G7A+E189P+E259G+N279S+K280Y+G303R、G7A+E18 9P+E259G+N279S+K280Y+E390A、G7A+E189P+E259G+N279S+G303R+E390A、G7A+E189P+E259G+K280Y+G303R+E390A、G7A+E189P+N279S+K280Y+G303R+ E390A, G7A+Y242F+E259G+N279S+K280Y+G303R, G7A+Y242F+E259G+N279S+K280Y+E390A, G7A+Y242F+E259G+N279S+G303R+E390A, G7A+Y242F+E259G +K280Y+G303R+E390A、G7A+Y242F+N279S+K280Y+G303R+E390A、G7A+E259G+N279S+K280Y+G303R+E390A、W140Y+E189P+Y242F+E259G+N279S+K280Y、W140Y+E189P+Y242F+E259G+N279S+G303R, W140Y+E189P+Y242F+E259G+N2 79S+E390A, W140Y+E189P+Y242F+E259G+K280Y+G303R, W140Y+E189P+Y242F +E259G+K280Y+E390A, W140Y+E189P+Y242F+E259G+G303R+E390A, W140Y+E 189P+Y242F+N279S+K280Y+G303R, W140Y+E189P+Y242F+N279S+K280Y+E390 A, W140Y+E189P+Y242F+N279S+G303R+E390A, W140Y+E189P+Y242F+K280Y+ G303R+E390A, W140Y+E189P+E259G+N279S+K280Y+G303R, W140Y+E189P+E25 9G+N279S+K280Y+E390A, W140Y+E189P+E259G+N279S+G303R+E390A, W140Y +E189P+E259G+K280Y+G303R+E390A, W140Y+E189P+N279S+K280Y+G303R+E3 90A, W140Y+Y242F+E259G+N279S+K280Y+G303R, W140Y+Y242F+E259G+N279 S+K280Y+E390A, W140Y+Y242F+E259G+N279S+G303R+E390A, W140Y+Y242F+E 259G+K280Y+G303R+E390A, W140Y+Y242F+N279S+K280Y+G303R+E390A, W14 0Y+E259G+N279S+K280Y+G303R+E390A, E189P+Y242F+E259G+N279S+K280Y+ The variant includes modifications at positions corresponding to G303R, E189P+Y242F+E259G+N279S+K280Y+E390A, E189P+Y242F+E259G+N279S+G303R+E390A, E189P+Y242F+E259G+K280Y+G303R+E390A, E189P+Y242F+N279S+K280Y+G303R+E390A, and the variant includes modifications at positions corresponding to SEQ ID NO: 1,The mutant has at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%, and the mutant has α-amylase activity.

[0380] In one embodiment, the α-amylase mutant of SEQ ID NO: 1 is a) Using sequence number 1 for numbering, deletes are made at positions corresponding to R180* and S181*, b) Using sequence number 1 for numbering, H1*+G7A+W140Y+E189P+Y242F+E259G+N279S, H1*+G7A+W140Y+E189P+Y242F+E259G+K280Y, H1*+G7A+W140Y+E189P+Y242F+E259G+G303R, H1*+G7A+W140Y+E189P+Y242F+E259G+E390A, H1*+G7A+W140Y+E189P+Y242F+N279S+K280Y, H1*+G7A+W140Y+E189P+Y242F+N279S+G303R, H1*+G7A+W140Y+E189P+Y242F+N279S+E390A, H1*+G7A+W140Y+E189P+Y242F+K280Y+G303R, H1*+G7A+W140Y+E189P+Y242F+K280Y+E390A, H1*+G7A+W140Y+E189P+Y242F+G303R+E390A, H1*+G7A+W140Y+E189P+E259G+N279S+K280Y, H1*+G7A+W140Y+E189P+E259G+N279S+G303R、 H1*+G7A+W140Y+E189P+E259G+N279S+E390A、 H1*+G7A+W140Y+E189P+E259G+K280Y+G303R、 H1*+G7A+W140Y+E189P+E259G+K280Y+E390A、 H1*+G7A+W140Y+E189P+E259G+G303R+E390A、 H1*+G7A+W140Y+E189P+N279S+K280Y+G303R、 H1*+G7A+W140Y+E189P+N279S+K280Y+E390A、 H1*+G7A+W140Y+E189P+N279S+G303R+E390A、 H1*+G7A+W140Y+E189P+K280Y+G303R+E390A、 H1*+G7A+W140Y+Y242F+E259G+N279S+K280Y、 H1*+G7A+W140Y+Y242F+E259G+N279S+G303R、 H1*+G7A+W140Y+Y242F+E259G+N279S+E390A、 H1*+G7A+W140Y+Y242F+E259G+K280Y+G303R、 H1*+G7A+W140Y+Y242F+E259G+K280Y+E390A、 H1*+G7A+W140Y+Y242F+E259G+G303R+E390A、 H1*+G7A+W140Y+Y242F+N279S+K280Y+G303R、 H1*+G7A+W140Y+Y242F+N279S+K280Y+E390A、 H1*+G7A+W140Y+Y242F+N279S+G303R+E390A、 H1*+G7A+W140Y+Y242F+K280Y+G303R+E390A、 H1*+G7A+W140Y+E259G+N279S+K280Y+G303Rぁ H1*+G7A+W140Y+E259G+N279S+K280Y+E390Aぁ H1*+G7A+W140Y+E259G+N279S+G303R+E390Aぁ H1*+G7A+W140Y+E259G+K280Y+G303R+E390A、 H1*+G7A+W140Y+N279S+K280Y+G303R+E390Aぁ H1*+G7A+E189P+Y242F+E259G+N279S+K280Yぁ H1*+G7A+E189P+Y242F+E259G+N279S+G303Rぁ H1*+G7A+E189P+Y242F+E259G+N279S+E390Aぁ H1*+G7A+E189P+Y242F+E259G+K280Y+G303Rぁ H1*+G7A+E189P+Y242F+E259G+K280Y+E390Aぁ H1*+G7A+E189P+Y242F+E259G+G303R+E390Aぁ H1*+G7A+E189P+Y242F+N279S+K280Y+G303Rぁ H1*+G7A+E189P+Y242F+N279S+K280Y+E390Aぁ H1*+G7A+E189P+Y242F+N279S+G303R+E390Aぁ H1*+G7A+E189P+Y242F+K280Y+G303R+E390Aぁ H1*+G7A+E189P+E259G+N279S+K280Y+G303Rぁ H1*+G7A+E189P+E259G+N279S+K280Y+E390Aぁ H1*+G7A+E189P+E259G+N279S+G303R+E390Aぁ H1*+G7A+E189P+E259G+K280Y+G303R+E390Aぁ H1*+G7A+E189P+N279S+K280Y+G303R+E390A、 H1*+G7A+Y242F+E259G+N279S+K280Y+G303R、 H1*+G7A+Y242F+E259G+N279S+K280Y+E390A、 H1*+G7A+Y242F+E259G+N279S+G303R+E390A、 H1*+G7A+Y242F+E259G+K280Y+G303R+E390A、 H1*+G7A+Y242F+N279S+K280Y+G303R+E390A、 H1*+G7A+E259G+N279S+K280Y+G303R+E390A、 H1*+W140Y+E189P+Y242F+E259G+N279S+K280Y、 H1*+W140Y+E189P+Y242F+E259G+N279S+G303R、 H1*+W140Y+E189P+Y242F+E259G+N279S+E390A、 H1*+W140Y+E189P+Y242F+E259G+K280Y+G303R、 H1*+W140Y+E189P+Y242F+E259G+K280Y+E390A、 H1*+W140Y+E189P+Y242F+E259G+G303R+E390A、 H1*+W140Y+E189P+Y242F+N279S+K280Y+G303R、 H1*+W140Y+E189P+Y242F+N279S+K280Y+E390A、 H1*+W140Y+E189P+Y242F+N279S+G303R+E390A、 H1*+W140Y+E189P+Y242F+K280Y+G303R+E390A、 H1*+W140Y+E189P+E259G+N279S+K280Y+G303R、 H1*+W140Y+E189P+E259G+N279S+K280Y+E390A、 H1*+W140Y+E189P+E259G+N279S+G303R+E390A、 H1*+W140Y+E189P+E259G+K280Y+G303R+E390A、 H1*+W140Y+E189P+N279S+K280Y+G303R+E390A、 H1*+W140Y+Y242F+E259G+N279S+K280Y+G303R、 H1*+W140Y+Y242F+E259G+N279S+K280Y+E390A、 H1*+W140Y+Y242F+E259G+N279S+G303R+E390A、 H1*+W140Y+Y242F+E259G+K280Y+G303R+E390A、 H1*+W140Y+Y242F+N279S+K280Y+G303R+E390A、 H1*+W140Y+E259G+N279S+K280Y+G303R+E390A、 H1*+E189P+Y242F+E259G+N279S+K280Y+G303R、 H1*+E189P+Y242F+E259G+N279S+K280Y+E390A、H1*+E189P+Y242F+E259G+N279S+G303R+E390A、 H1*+E189P+Y242F+E259G+K280Y+G303R+E390A、H1*+E189P+Y242F+N279S+K280Y+G303R+E390A、 H1*+E189P+E259G+N279S+K280Y+G303R+E390A、H1*+Y242F+E259G+N279S+K280Y+G303R+E390A、 G7A+W140Y+E189P+Y242F+E259G+N279S+K280Y、 G7A+W140Y+E189P+Y242F+E259G+N279S+G303R、 G7A+W140Y+E189P+Y242F+E259G+N279S+E390A、 G7A+W140Y+E189P+Y242F+E259G+K280Y+G303R、 G7A+W140Y+E189P+Y242F+E259G+K280Y+E390A、 G7A+W140Y+E189P+Y242F+E259G+G303R+E390Aぁ G7A+W140Y+E189P+Y242F+N279S+K280Y+G303R、 G7A+W140Y+E189P+Y242F+N279S+K280Y+E390A、 G7A+W140Y+E189P+Y242F+N279S+G303R+E390Aぁ G7A+W140Y+E189P+Y242F+K280Y+G303R+E390A、 G7A+W140Y+E189P+E259G+N279S+K280Y+G303Rぁ G7A+W140Y+E189P+E259G+N279S+K280Y+E390A、 G7A+W140Y+E189P+E259G+N279S+G303R+E390Aぁ G7A+W140Y+E189P+E259G+K280Y+G303R+E390A、 G7A+W140Y+E189P+N279S+K280Y+G303R+E390A、 G7A+W140Y+Y242F+E259G+N279S+K280Y+G303Rぁ G7A+W140Y+Y242F+E259G+N279S+K280Y+E390Aぁ G7A+W140Y+Y242F+E259G+N279S+G303R+E390Aぁ G7A+W140Y+Y242F+E259G+K280Y+G303R+E390Aぁ G7A+W140Y+Y242F+N279S+K280Y+G303R+E390Aぁ G7A+W140Y+E259G+N279S+K280Y+G303R+E390Aぁ G7A+E189P+Y242F+E259G+N279S+K280Y+G303Rぁ G7A+E189P+Y242F+E259G+N279S+K280Y+E390Aぁ G7A+E189P+Y242F+E259G+N279S+G303R+E390Aぁ G7A+E189P+Y242F+E259G+K280Y+G303R+E390Aぁ G7A+E189P+Y242F+N279S+K280Y+G303R+E390Aぁ G7A+E189P+E259G+N279S+K280Y+G303R+E390Aぁ G7A+Y242F+E259G+N279S+K280Y+G303R+E390Aぁ W140Y+E189P+Y242F+E259G+N279S+K280Y+G303R、 W140Y+E189P+Y242F+E259G+N279S+K280Y+E390A、 W140Y+E189P+Y242F+E259G+N279S+G303R+E390A、 W140Y+E189P+Y242F+E259G+K280Y+G303R+E390A、 W140Y+E189P+Y242F+N279S+K280Y+G303R+E390A、 W140Y+E189P+E259G+N279S+K280Y+G303R+E390A、 The variant includes modifications at positions corresponding to W140Y+Y242F+E259G+N279S+K280Y+G303R+E390A and E189P+Y242F+E259G+N279S+K280Y+G303R+E390A, wherein the variant is at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, and less than 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%. At least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, provided that the mutant has less than 100% sequence homology and the mutant has α-amylase activity.

[0381] In one embodiment, the α-amylase mutant of SEQ ID NO: 1 is a) Using sequence number 1 for numbering, deletes are made at positions corresponding to R180* and S181*, b) Using sequence number 1 for numbering: H1*+G7A+W140Y+E189P+Y242F+E259G+N279S+K280Y, H1*+G7A+W140Y+E189P+Y242F+E259G+N279S+G303R, H1*+G7A+W140Y+E189P+Y242F+E259G+N279S+E390A, H1*+G7A+W140Y+E189P+Y242F+E259G+K280Y+G303R, H1*+G7A+W140Y+E189P+Y242F+E259G+K280Y+E390A, H1*+G7A +W140Y+E189P+Y242F+E259G+G303R+E390A, H1*+G7A+W140Y+E189P+Y242 F+N279S+K280Y+G303R, H1*+G7A+W140Y+E189P+Y242F+N279S+K280Y+E390 A, H1*+G7A+W140Y+E189P+Y242F+N279S+G303R+E390A, H1*+G7A+W140Y+E1 89P+Y242F+K280Y+G303R+E390A, H1*+G7A+W140Y+E189P+E259G+N279S+K2 80Y+G303R, H1*+G7A+W140Y+E189P+E259G+N279S+K280Y+E390A, H1*+G7A +W140Y+E189P+E259G+N279S+G303R+E390A, H1*+G7A+W140Y+E189P+E259G +K280Y+G303R+E390A, H1*+G7A+W140Y+E189P+N279S+K280Y+G303R+E390A , H1*+G7A+W140Y+Y242F+E259G+N279S+K280Y+G303R, H1*+G7A+W140Y+Y24 2F+E259G+N279S+K280Y+E390A, H1*+G7A+W140Y+Y242F+E259G+N279S+G3 03R+E390A, H1*+G7A+W140Y+Y242F+E259G+K280Y+G303R+E390A, H1*+G7A+ W140Y+Y242F+N279S+K280Y+G303R+E390A, H1*+G7A+W140Y+E259G+N279S+ K280Y+G303R+E390A, H1*+G7A+E189P+Y242F+E259G+N279S+K280Y+G303R,H1*+G7A+E189P+Y242F+E259G+N279S+K280Y+E390A、H1*+G7A+E189P+Y242F+E259G+N279S+G303R+E390A、H1*+G7A+E189P+Y242F+E259G+K280Y+G303R +E390A, H1*+G7A+E189P+Y242F+N279S+K280Y+G303R+E390A, H1*+G7A+E189P+E259G+N279S+K280Y+G303R+E390A, H1*+G7A+Y242F+E259G+N279S+K280 Y+G303R+E390A、H1*+W140Y+E189P+Y242F+E259G+N279S+K280Y+G303R、H1*+W140Y+E189P+Y242F+E259G+N279S+K280Y+E390A、H1*+W140Y+E189P+Y24 2F+E259G+N279S+G303R+E390A、H1*+W140Y+E189P+Y242F+E259G+K280Y+G 303R+E390A、H1*+W140Y+E189P+Y242F+N279S+K280Y+G303R+E390A、H1*+W1 40Y+E189P+E259G+N279S+K280Y+G303R+E390A、H1*+W140Y+Y242F+E259G+ N279S+K280Y+G303R+E390A、H1*+E189P+Y242F+E259G+N279S+K280Y+G303R +E390A, G7A+W140Y+E189P+Y242F+E259G+N279S+K280Y+G303R, G7A+W140Y+E189P+Y242F+E259G+N279S+K280Y+E390A, G7A+W140Y+E189P+Y242F+E259 G+N279S+G303R+E390A、G7A+W140Y+E189P+Y242F+E259G+K280Y+G303R+E3 90A、G7A+W140Y+E189P+Y242F+N279S+K280Y+G303R+E390A、G7A+W140Y+E18 9P+E259G+N279S+K280Y+G303R+E390A、G7A+W140Y+Y242F+E259G+N279S+K280Y+G303R+E390A、G7A+E189P+Y242F+E259G+N279S+K280Y+G303R+E390A、The variant includes a modification at the position corresponding to W140Y+E189P+Y242F+E259G+N279S+K280Y+G303R+E390A, wherein the variant has at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%, and the variant has α-amylase activity.

[0382] In one embodiment, the α-amylase mutant of SEQ ID NO: 1 is a) Using sequence number 1 for numbering, deletes are made at positions corresponding to R180* and S181*, b) Using sequence number 1 for numbering, H1*+G7A+W140Y+E189P+Y242F+E259G+N279S+K280Y+G303R, H1*+G7A+W140Y+E189P+Y242F+E259G+N279S+K280Y+E390A, H1*+G7A+W140Y+E189P+Y242F+E259G+N279S+G303R+E390A, H1*+G7A+W140Y+E189P+Y242F+E259G+K280Y+G3 03R+E390A, H1*+G7A+W140Y+E189P+Y242F+N279S+K280Y+G303R+E390A, H1*+G7A+W140Y+E189P+E259G+N279S+K280Y+G303R+E 390A, H1*+G7A+W140Y+Y242F+E259G+N279S+K280Y+G303R+E390A, H1*+G7A+E189P+Y242F+E259G+N279S+K280Y+G303R+E390A, The variant includes modifications at positions corresponding to H1*+W140Y+E189P+Y242F+E259G+N279S+K280Y+G303R+E390A and G7A+W140Y+E189P+Y242F+E259G+N279S+K280Y+G303R+E390A, wherein the variant comprises at least 60%, at least 65%, at least 70%, at least 75%, and at least 80% of sequence numbers 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14, At least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, provided that the sequence homology is less than 100%, and the mutant has α-amylase activity.

[0383] In one embodiment, the α-amylase mutant of SEQ ID NO: 1 is a) Using sequence number 1 for numbering, deletes are made at positions corresponding to R180* and S181*, The variant includes modifications at positions corresponding to H1*+G7A+W140Y+E189P+Y242F+E259G+N279S+K280Y+G303R+E390A using sequence number 1 for numbering, wherein the variant is at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, and at least 8 2%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, provided that the sequence homology is less than 100%, and the mutant has α-amylase activity.

[0384] In one embodiment, the α-amylase mutant of SEQ ID NO: 2 is a) Using sequence number 1 for numbering, deletes are made at positions corresponding to R180* and S181*, b) Use array index 1 for numbering: H1*+D118T, H1*+W140Y, H1*+T182S, H1*+A185D, H1*+E189P, H1*+Y242F, H1*+E259G, H1*+F266Y, H1*+N279S, H1*+K280Y, H1*+G303R, H1*+E390A, H1*+G475K, H1*+A185Q, H1*+N194F, D118T+W140Y, D118T+T182S, D118T+A185D, D118T+E189P, D118T+Y242F, D118T+E259G, D118T+F2 66Y, D118T+N279S, D118T+K280Y, D118T+G303R, D118T+E390A, D118T+G475 K, D118T+A185Q, D118T+N194F, W140Y+T182S, W140Y+A185D, W140Y+E189P, W 140Y+Y242F, W140Y+E259G, W140Y+F266Y, W140Y+N279S, W140Y+K280Y, W14 0Y+G303R, W140Y+E390A, W140Y+G475K, W140Y+A185Q, W140Y+N194F, T182S+ A185D, T182S+E189P, T182S+Y242F, T182S+E259G, T182S+F266Y, T182S+N2 79S, T182S+K280Y, T182S+G303R, T182S+E390A, T182S+G475K, T182S+A185Q , T182S+N194F, A185D+E189P, A185D+Y242F, A185D+E259G, A185D+F266Y, A 185D+N279S, A185D+K280Y, A185D+G303R, A185D+E390A, A185D+G475K, A185 D, +A185Q, A185D, +N194F, E189P+Y242F, E189P+E259G, E189P+F266Y, E189 P+N279S, E189P+K280Y, E189P+G303R, E189P+E390A, E189P+G475K, E189P+A 185Q, E189P+N194F, Y242F+E259G, Y242F+F266Y, Y242F+N279S, Y242F+K28 0Y, Y242F+G303R, Y242F+E390A, Y242F+G475K, Y242F+A185Q, Y242F+N194F,E259G+F266Y, E259G+N279S, E259G+K280Y, E259G+G303R, E259G+E390A, E259G+G475K, E259G+A185Q, E259G+N19 4F, F266Y+N279S, F266Y+K280Y, F266Y+G303R, F266Y+E390A, F266Y+G475K, F266Y+A185Q, F266Y+N194F, N279+K 280Y, N279S+G303R, N279S+E390A, N279S+G475K, N279S+A185Q, N279S+N194F, K280Y+G303R, K280Y+E390A, K280 Y+G475K, K280Y+A185Q, K280Y+N194F, G303R+E390A, G303R+G475K, G303R+A185Q, G303R+N194F, E390A+G475K, E3 The variant includes modifications at positions corresponding to 90A+A185Q, E390A+N194F, G475K+A185Q, G475K+N194F, and A185Q+N194F, wherein the variant is at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, and at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, provided that the sequence homology is less than 100%, and the variant has α-amylase activity.

[0385] In one embodiment, the α-amylase mutant of SEQ ID NO: 2 is a) Using sequence number 1 for numbering, deletes are made at positions corresponding to R180* and S181*, b) Using array index 1 for numbering: H1*+D118T+W140Y, H1*+D118T+T182S, H1*+D118T+A185D, H1*+D118T+E189P, H1*+D118T+Y242F, H1*+D118T+E259G, H1*+D118T+F266Y, H1*+D118T+N279S, H1*+D118T+K280Y, H1*+D118T+G303R, H1*+D118T+E390A, H1*+D118T+G475K, H1*+D118T+A185Q, H1*+D118T+N194F, H1*+W 140Y+T182S, H1*+W140Y+A185D, H1*+W140Y+E189P, H1*+W140Y+Y242F, H1* +W140Y+E259G, H1*+W140Y+F266Y, H1*+W140Y+N279S, H1*+W140Y+K280Y, H 1*+W140Y+G303R, H1*+W140Y+E390A, H1*+W140Y+G475K, H1*+W140Y+A185Q , H1*+W140Y+N194F, H1*+T182S+A185D, H1*+T182S+E189P, H1*+T182S+Y242 F, H1*+T182S+E259G, H1*+T182S+F266Y, H1*+T182S+N279S, H1*+T182S+K2 80Y, H1*+T182S+G303R, H1*+T182S+E390A, H1*+T182S+G475K, H1*+T182S+ A185Q, H1*+T182S+N194F, H1*+A185D+E189P, H1*+A185D+Y242F, H1*+A185 D+E259G, H1*+A185D+F266Y, H1*+A185D+N279S, H1*+A185D+K280Y, H1*+A18 5D+G303R, H1*+A185D+E390A, H1*+A185D+G475K, H1*+A185D+A185Q, H1*+A 185D+N194F, H1*+E189P+Y242F, H1*+E189P+E259G, H1*+E189P+F266Y, H1* +E189P+N279S, H1*+E189P+K280Y, H1*+E189P+G303R, H1*+E189P+E390A, H 1*+E189P+G475K, H1*+E189P+A185Q, H1*+E189P+N194F, H1*+Y242F+E259G,H1*+Y242F+F266Y、H1*+Y242F+N279S、H1*+Y242F+K280Y、H1*+Y242F+G303R、H1*+Y242F+E390A、H1*+Y242F+G475K、H1*+Y242F+A185Q、H1*+Y242F+N194F、H1*+E259G+F266Y、H1*+E259G+N279S、H1*+E259G+K280Y、H1*+E259G+G303R、H1*+E259G+E390A、H1*+E259G+G475K、H1*+E259G+A185Q、H1*+E259G+N194F、H1*+F266Y+N279S、H1*+F266Y+K280Y、H1*+F266Y+G303R、H1*+F266Y+E390A、H1*+F266Y+G475K、H1*+F266Y+A185Q、H1*+F266Y+N194F、H1*+N279S+K280Y、H1*+N279S+G303R、H1*+N279S+E390A、H1*+N279S+G475K、H1*+N279S+A185Q、H1*+N279S+N194F、H1*+K280Y+G303R、H1*+K280Y+E390A、H1*+K280Y+G475K、H1*+K280Y+A185Q、H1*+K280Y+N194F、H1*+G303R+E390A、H1*+G303R+G475K、H1*+G303R+A185Q、H1*+G303R+N194F、H1*+E390A+G475K、H1*+E390A+A185Q、H1*+E390A+N194F、H1*+G475K+A185Q、H1*+G475K+N194F、H1*+A185Q+N194F、D118T+W140Y+T182S、D118T+W140Y+A185D、D118T+W140Y+E189P、D118T+W140Y+Y242F、D118T+W140Y+E259G、D118T+W140Y+F266Y、D118T+W140Y+N279S、D118T+W140Y+K280Y、D118T+W140Y+G303R、D118T+W140Y+E390A、D118T+W140Y+G475K、D118T+W140Y+A185Q、D118T+W140Y+N194F、D118T+T182S+A185D、D118T+T182S+E189P、D118T+T182S+Y242F、D118T+T182S+E259G、D118T+T182S+F266Y、D118T+T182S+N279S、D118T+T182S+K280Y、D118T+T182S+G303R、D118T+T182S+E390A、D118T+T182S+G475K、D118T+T182S+A185Q、D118T+T182S+N194F、D118T+A185D+E189P、D118T+A185D+Y242F、D118T+A185D+E259G、D118T+A185D+F266Y、D118T+A185D+N279S、D118T+A185D+K280Y、D118T+A185D+G303R、D118T+A185D+E390A、D118T+A185D+G475K、D118T+A185D+A185Q、D118T+A185D+N194F、D118T+E189P+Y242F、D118T+E189P+E259G、D118T+E189P+F266Y、D118T+E189P+N279S、D118T+E189P+K280Y、D118T+E189P+G303R、D118T+E189P+E390A、D118T+E189P+G475K、D118T+E189P+A185Q、D118T+E189P+N194F、D118T+Y242F+E259G、D118T+Y242F+F266Y、D118T+Y242F+N279S、D118T+Y242F+K280Y、D118T+Y242F+G303R、D118T+Y242F+E390A、D118T+Y242F+G475K、D118T+Y242F+A185Q、D118T+Y242F+N194F、D118T+E259G+F266Y、D118T+E259G+N279S、D118T+E259G+K280Y、D118T+E259G+G303R、D118T+E259G+E390A、D118T+E259G+G475K、D118T+E259G+A185Q、D118T+E259G+N194F、D118T+F266Y+N279S、D118T+F266Y+K280Y、D118T+F266Y+G303R、D118T+F266Y+E390A、D118T+F266Y+G475K、D118T+F266Y+A185Q、D118T+F266Y+N194F、D118T+N279S+K280Y、D118T+N279S+G303R、D118T+N279S+E390A、D118T+N279S+G475K、D118T+N279S+A185Q、D118T+N279S+N194F、D118T+K280Y+G303R、D118T+K280Y+E390A、D118T+K280Y+G475K、D118T+K280Y+A185Q、D118T+K280Y+N194F、D118T+G303R+E390A、D118T+G303R+G475K、D118T+G303R+A185Q、D118T+G303R+N194F、D118T+E390A+G475K、D118T+E390A+A185Q、D118T+E390A+N194F、D118T+G475K+A185Q、D118T+G475K+N194F、D118T+A185Q+N194F、W140Y+T182S+A185D、W140Y+T182S+E189P、W140Y+T182S+Y242F、W140Y+T182S+E259G、W140Y+T182S+F266Y、W140Y+T182S+N279S、W140Y+T182S+K280Y、W140Y+T182S+G303R、W140Y+T182S+E390A、W140Y+T182S+G475K、W140Y+T182S+A185Q、W140Y+T182S+N194F、W140Y+A185D+E189P、W140Y+A185D+Y242F、W140Y+A185D+E259G、W140Y+A185D+F266Y、W140Y+A185D+N279S、W140Y+A185D+K280Y、W140Y+A185D+G303R、W140Y+A185D+E390A、W140Y+A185D+G475K、W140Y+A185D+A185Q、W140Y+A185D+N194F、W140Y+E189P+Y242F、W140Y+E189P+E259G、W140Y+E189P+F266Y、W140Y+E189P+N279S、W140Y+E189P+K280Y、W140Y+E189P+G303R、W140Y+E189P+E390A、W140Y+E189P+G475K、W140Y+E189P+A185Q、W140Y+E189P+N194F、W140Y+Y242F+E259G、W140Y+Y242F+F266Y、W140Y+Y242F+N279S、W140Y+Y242F+K280Y、W140Y+Y242F+G303R、W140Y+Y242F+E390A、W140Y+Y242F+G475K、W140Y+Y242F+A185Q、W140Y+Y242F+N194F、W140Y+E259G+F266Y、W140Y+E259G+N279S、W140Y+E259G+K280Y、W140Y+E259G+G303R、W140Y+E259G+E390A、W140Y+E259G+G475K、W140Y+E259G+A185Q、W140Y+E259G+N194F、W140Y+F266Y+N279S、W140Y+F266Y+K280Y、W140Y+F266Y+G303R、W140Y+F266Y+E390A、W140Y+F266Y+G475K、W140Y+F266Y+A185Q、W140Y+F266Y+N194F、W140Y+N279S+K280Y、W140Y+N279S+G303R、W140Y+N279S+E390A、W140Y+N279S+G475K、W140Y+N279S+A185Q、W140Y+N279S+N194F、W140Y+K280Y+G303R、W140Y+K280Y+E390A、W140Y+K280Y+G475K、W140Y+K280Y+A185Q、W140Y+K280Y+N194F、W140Y+G303R+E390A、W140Y+G303R+G475K、W140Y+G303R+A185Q、W140Y+G303R+N194F、W140Y+E390A+G475K、W140Y+E390A+A185Q、W140Y+E390A+N194F、W140Y+G475K+A185Q、W140Y+G475K+N194F、W140Y+A185Q+N194F、T182S+A185D+E189P、T182S+A185D+Y242F、T182S+A185D+E259G、T182S+A185D+F266Y、T182S+A185D+N279S、T182S+A185D+K280Y、T182S+A185D+G303R、T182S+A185D+E390A、T182S+A185D+G45K、T182S+A185D+A185Q、T182S+A185D+N194F、T182S+E189P+Y242F、T182S+E189P+E259G、T182S+E189P+F266Y、T182S+E189P+N279S、T182S+E189P+K280Y、T182S+E189P+G303R、T182S+E189P+E390A、T182S+E189P+G475K、T182S+E189P+A185Q、T182S+E189P+N194F、T1、 82S+Y242F+E259G、T182S+Y242F+F266Y、T182S+Y242F+N279S、T182S+Y242F+K280Y、T182S+Y242F+G303R、T182S+Y242F+E390A、T182S+Y242F+G475K、T182S+Y242F+A185Q、T182S+Y242F+N194F、T182S+E259G+F266Y、T182S+E259G+N279S、T182S+E259G+K280Y、T182S+E259G+G303R、T182S+E259G+E390A、T182S+E259G+G475K、T182S+E259G+A185Q、T182S+E259G+N194F、T182S+F266Y+N279S、T182S+F266Y+K280Y、T182S+F266Y+G303R、T182S+F266Y+E390A、T182S+F266Y+G475K、T182S+F266Y+A185Q、T182S+F266Y+N194F、T182S+N279S+K280Y、T182S+N279S+G303R、T182S+N279S+E390A、T182S+N279S+G475K、T182S+N279S+A185Q、T182S+N279S+N194F、T182S+K280Y+G303R、T182S+K280Y+E390A、T182S+K280Y+G475K、T182S+K280Y+A185Q、T182S+K280Y+N194F、T182S+G303R+E390A、T182S+G303R+G475K、T182S+G303R+A185Q、T182S+G303R+N194F、T182S+E390A+G475K、T182S+E390A+A185Q、T182S+E390A+N194F、T182S+G475K+A185Q、T182S+G475K+N194F、T182S+A185Q+N194F、A185D+E189P+Y242F、A185D+E189P+E259G、A185D+E189P+F266Y、A185D+E189P+N279S、A185D+E189P+K280Y、A185D+E189P+G303R、A185D+E189P+E390A、A185D+E189P+G475K、A185D+E189P+A185Q、A185D+E189P+N194F、A185D+Y242F+E259G、A185D+Y242F+F266Y、A185D+Y242F+N279S、A185D+Y242F+K280Y、A185D+Y242F+G303R、A185D+Y242F+E390A、A185D+Y242F+G475K、A185D+Y242F+A185Q、A185D+Y242F+N194F、A185D+E259G+F266Y、A185D+E259G+N279S、A185D+E259G+K280Y、A185D+E259G+G303R、A185D+E259G+E390A、A185D+E259G+G475K、A185D+E259G+A185Q、A185D+E259G+N194F、A185D+F266Y+N279S、A185D+F266Y+K280Y、A185D+F266Y+G303R、A185D+F266Y+E390A、A185D+F266Y+G475K、A185D+F266Y+A185Q、A185D+F266Y+N194F、A185D+N279S+K280Y、A185D+N279S+G303R、A185D+N279S+E390A、A185D+N279S+G475K、A185D+N279S+A185Q、A185D+N279S+N194F、A185D+K280Y+G303R、A185D+K280Y+E390A、A185D+K280Y+G475K、A185D+K280Y+A185Q、A185D+K280Y+N194F、A185D+G303R+E390A、A185D+G303R+G475K、A185D+G303R+A185Q、A185D+G303R+N194F、A185D+E390A+G475K、A185D+E390A+A185Q、A185D+E390A+N194F、A185D+G475K+A185Q、A185D+G475K+N194F、A185D+A185Q+N194F、E189P+Y242F+E259G、E189P+Y242F+F266Y、E189P+Y242F+N279S、E189P+Y242F+K280Y、E189P+Y242F+G303R、E189P+Y242F+E390A、E189P+Y242F+G475K、E189P+Y242F+A185Q、E189P+Y242F+N194F、E189P+E259G+F266Y、E189P+E259G+N279S、E189P+E259G+K280Y、E189P+E259G+G303R、E189P+E259G+E390A、E189P+E259G+G475K、E189P+E259G+A185Q、E189P+E259G+N194F、E189P+F266Y+N279S、E189P+F266Y+K280Y、E189P+F266Y+G303R、E189P+F266Y+E390A、E189P+F266Y+G475K、E189P+F266Y+A185Q、E189P+F266Y+N194F、E189P+N279S+K280Y、E189P+N279S+G303R、E189P+N279S+E390A、E189P+N279S+G475K、E189P+N279S+A185Q、E189P+N279S+N194F、E189P+K280Y+G303R、E189P+K280Y+E390A、E189P+K280Y+G475K、E189P+K280Y+A185Q、E189P+K280Y+N194F、E189P+G303R+E390A、E189P+G303R+G475K、E189P+G303R+A185Q、E189P+G303R+N194F、E189P+E390A+G475K、E189P+E390A+A185Q、E189P+E390A+N194F、E189P+G475K+A185Q、E189P+G475K+N194F、E189P+A185Q+N194F、Y242F+E259G+F266Y、Y242F+E259G+N279S、Y242F+E259G+K280Y、Y242F+E259G+G303R、Y242F+E259G+E390A、Y242F+E259G+G475K、Y242F+E259G+A185Q、Y242F+E259G+N194F、Y242F+F266Y+N279S、Y242F+F266Y+K280Y、Y242F+F266Y+G303R、Y242F+F266Y+E390A、Y242F+F266Y+G475K、Y242F+F266Y+A185Q、Y242F+F266Y+N194F、Y242F+N279S+K280Y、Y242F+N279S+G303R、Y242F+N279S+E390A、Y242F+N279S+G475K、Y242F+N279S+A185Q、Y242F+N279S+N194F、Y242F+K280Y+G303R、Y242F+K280Y+E390A、Y242F+K 280Y+G475K、Y242F+K280Y+A185Q、Y242F+K280Y+N194F、Y242F+G303R+E39 0A、Y242F+G303R+G475K、Y242F+G303R+A185Q、Y242F+G303R+N194F、Y242F +E390A+G475K、Y242F+E390A+A185Q、Y242F+E390A+N194F、Y242F+G475K+A 185Q、Y242F+G475K+N194F、Y242F+A1...

Claims

1. 10. An α-amylase variant of a parent α-amylase, said variant comprising a substitution corresponding to G303R, using SEQ ID NO: 1 for numbering, as well as H1*, H2*, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F116L, F116N, F116Q, F116W, V117G, D1 18Q, D118T, D123N, N126D, N128Y, T131I, T131V, S132D, S132L, T134D, Y135E , Q136T, Q136K, I137F, Q138V, W140S, W140M, W140Y, K142E, K142G, K142P, D 144N, P146S, P146H, T151A, Y152L, F155W, W159H, W159L, Y160P, T165V, W167 A, W167D, W167E, W167F, W167P, W167S, E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K178A, R180*, R180Q, R180S, S181*, S18 1L, S181T, T182*, T182D, T182E, T182G, T182I, T182L, T182M, T182S, G183* , G183P, A185D, A185E, A185K, A185N, A185Q, A185R, W188C, E189P, E193I, N1 94F, F202Y, F202Y, A203F, L205F, L205N, L205Q, L205R, L205V, L205Y, T224 F, T224Y, N225S, Y242F, Y242L, S243Q, F244I, F244L, V252I, T256A, T256D, K 258V, E259G, E259V, M260D, M260W, F261K, F261P, F261S, F266Y, N276F, N27 6W, N276Y, N279S, N280F, N280W, N280Y, W283N, S286K, S286L, S302T, S302Y,T315A, H320T, P321H, P321Y, M322G, M322Y, H323A, H323N, H323W, W346N, L354V, E3 59W, Q360M, Q360W, Q360Y, G361M, Y362V, V365I, Y371S, Y371N, A383D, E390A, E390 D, N394W, N394D, F395Y, D405G, I409L, R414M, H420P, H420A, H420Q, H420Y, N422V, T427L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G47 α-amylase variants comprising one or more further substitutions or deletions corresponding to G5E, G475K, G476H, G476E and G476A, where "*" denotes a deletion, wherein the variant has at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%, sequence identity with SEQ ID NO: 1, and wherein the variant has improved cleaning performance as measured by specific activity of an improvement factor (IF) > 1.0 when compared to the parent α-amylase in SEQ ID NO:

2.

2. 2. The mutant of claim 1, wherein the mutant comprises deletions at two positions selected from the group consisting of R180* + S181*; R180* + T182*; R180* + G183*; S181* + T182*; S181* + G183*; and T182* + G183*, using SEQ ID NO: 1 for numbering.

3. 3. The mutant of claim 1 or 2, wherein the mutant is (a) substitutions and / or deletions at two, three, or four positions in said parent α-amylase, said positions corresponding to positions 180, 181, 182, and 183 using SEQ ID NO: 1 for numbering; (b) Substitutions corresponding to G303R, as well as H1*, H2, N3D, N3G, N3S, G7A, G7H, G7S, R26Y, A31S, R37A, R37N, R37V, T40G, T40N, A41D, N54S, V56T, A60V, A60P, L63F, K72R, G73L, K93R, N94D, Q98L, V103A, V103I, V103L, V104M, M105I, M105L, N106D, A109G, G110K, A111G, A111L, G113Q, G113S, G113A, T114K, F116L, F116N, F116Q, F116W, V117G, D118Q, D118T, D123N, N126D, N128Y, T1 31I, T131V, S132D, S132L, T134D, Y135E, Q136T, Q136K, I137F, Q138V, W140S , W140M, W140Y, K142E, K142G, K142P, D144N, P146S, P146H, T151A, Y152L, F 155W, W159H, W159L, Y160P, T165V, W167A, W167D, W167E, W167F, W167P, W167 S, E169I, E169Q, E169V, K172G, K172Q, L173F, N174K, N174S, I176V, Y177D, K178A, A185D, A185E, A185K, A185N, A185Q, A185R, W188C, E189P, E193I, N19 4F, F202Y, F202Y, A203F, L205F, L205N, L205Q, L205R, L205V, L205Y, T224F , T224Y, N225S, Y242F, Y242L, S243Q, F244I, F244L, V252I, T256A, T256D, K2 58V, E259G, E259V, M260D, M260W, F261K, F261P, F261S, F266Y, N276F, N276 W, N276Y, N279S, N280F, N280W, N280Y, W283N, S286K, S286L, S302T, S302Y, T 315A, H320T, P321H, P321Y, M322G, M322Y, H323A, H323N, H323W, W346N, L35 4V, E359W, Q360M, Q360W, Q360Y, G361M, Y362V, V365I, Y371S, Y371N, A383D,E390A, E390D, N394W, N394D, F395Y, D405G, I409L, R414M, H420P, H420A, H420Q, H420Y, N422V, T42 7L, G455R, K457N, K457R, R458S, N470A, N470M, S472R, V473C, N474F, N474L, G475E, G475K, G476H, G and one or more further substitutions or deletions corresponding to G476E and G476A, wherein the variant has at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%, sequence identity with SEQ ID NO:

1.

4. The mutant according to any one of claims 1 to 3, wherein the mutant is Using SEQ ID NO: 1 for numbering, H1*+G7A+R180*+S181*+Y242F+N279S+G303R, H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R, H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+E390A, H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K, H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G476A, H1*+G7A+R180*+S181*+D118T+Y242F+E259G+N279S+G303R+E390A+G475K, H1*+G7A+R180*+S181*+D118T+W140Y+Y242F+E259G+N279S+G303R+E390A+G476A, H1*+G7A+R180*+S181*+D118T+W140Y+Y242F+E259G+N279S+G303R+E390A+G475K, H1*+G7A+R180*+S181*+D118T+W140Y+Y242F+E259G+N279S+G303R+E390A+G476A, H1*+G7A+R180*+S181*+D118T+W140Y+N194F+Y242F+E259G+N279S+G303R+E390A+G475K, H1*+G7A+R180*+S181*+D118T+W140Y+N194F+Y242F+E259G+N279S+G303R+E390A+G476A, H1*+G7A+R180*+S181*+D118T+W140Y+N194F+Y242F+E259G+K280Y+N279S+G303R+E390A+G475K, 1. The variant comprises one or more substitutions or deletions selected from the group consisting of: H1* + G7A + R180* + S181* + D118T + W140Y + N194F + Y242F + E259G + K280Y + N279S + G303R + E390A + G476A, wherein the variant has at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, but less than 100%, sequence identity to SEQ ID NO: 1, and wherein the variant has α-amylase activity.

5. The mutant according to any one of claims 1 to 4, Using SEQ ID NO: 1 for numbering, H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K, H1*+G7A+S181*+T182*+Y242F+E259G+N279S+G303R+E390A+G475K, H1*+G7A+R180*+S181*+T182S+Y242F+E259G+N279S+G303R+E390A+G475K, H1*+G7A+R180*+T182L+G183*+Y242F+E259G+N279S+G303R+E390A+G475K, H1*+G7A+S181L+T182*+G183*+Y242F+E259G+N279S+G303R+E390A+G475K, H1*+G7A+R180S+T182*+G183*+Y242F+E259G+N279S+G303R+E390A+G475K, H1*+G7A+A60P+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K, H1*+G7A+M105L+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K, H1*+G7A+M105I+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K, H1*+G7A+R180*+S181*+T182D+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+T182E+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+T182G+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+T182M+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+A185K+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+A185D+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+A185Q+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+A185N+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+A185R+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+A185E+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+L205F+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+S243Q+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+F244L+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+F244I+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+N194F+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+N196F+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+N194F+L205F+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+N196F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+Y242F+E259G+N276F+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+Y242F+E259G+N276Y+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+Y242F+E259G+N276W+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+Y242F+E259G+N279S+K280W+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+L205F+Y242F+E259G+N276F+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+L205F+Y242F+E259G+N276Y+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+L205F+Y242F+E259G+N276W+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+L205F+Y242F+E259G+N279S+K280F+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+L205F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+L205F+Y242F+E259G+N279S+K280W+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+L205Y+Y242F+E259G+N276F+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+L205Y+Y242F+E259G+N276Y+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+L205Y+Y242F+E259G+N276W+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+L205Y+Y242F+E259G+N279S+K280F+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+E189P+L205Y+Y242F+E259G+N279S+K280W+G303R+E390A+G475K、 H1*+G7A+G113S+F116W+D123N+S181*+T182*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+F116L+D123N+N126D+T131I+S181*+T182*+F202Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7S+T40N+Q98L+R180*+S181*+Y242F+E259G+N279S+G303R+Y362V+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+M322G+E359W+Q360W+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+M322G+Q360W+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+E359W+Q360W+E390A+G475K、 H1*+G7S+R180*+S181*+Y242F+E259G+F261S+N279S+G303R+Y362V+E390A+G475K、 H1*+G7A+T40N+Q98L+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7S+T40N+Q98L+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+F261S+N279S+S286K+G303R+Y362V+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+W283N+S286K+G303R+H320T+H323A+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+P321H+M322Y+Q360Y+E390A+G475K、 H1*+G7A+D123N+I176V+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+G113A+F116W+D123N+T131V+Q136K+E169Q+K172Q+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+G113S+F116W+D123N+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+F116L+D123N+N126D+T131I+R180*+S181*+F202Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+N106D+A109G+F116W+D123N+N126D+T131I+T165V+N174S+R180*+S181*+T182I+F202Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7S+T40N+Q98L+R180*+S181*+Y242F+E259G+N279S+G303R+H320T+E390A+G475K、 H1*+G7S+T40N+Q98L+R180*+S181*+Y242F+E259G+N279S+G303R+Y362V+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+M322G+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+M322G+Q360W+E390A+G475K、 H1*+G7A+T40N+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7S+Q98L+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7S+R180*+S181*+Y242F+E259G+F261K+N279S+G303R+E390A+G475K、 H1*+G7S+R180*+S181*+Y242F+E259G+N279S+G303R+Y362V+E390A+G475K、 H1*+G7A+T40N+Q98L+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+Y362V+E390A+G475K、 H1*+G7S+R180*+S181*+Y242F+E259G+F261S+N279S+G303R+E390A+G475K、 H1*+G7A+I137F+R180*+S181*+Y242F+E259G+F261S+N279S+G303R+Y362V+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+S286K+G303R+H323A+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+H320T+H323A+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+P321H+M322Y+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+P321H+Q360Y+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+M322Y+Q360Y+E390A+G475K、 H1*+G7S+T40N+R180*+S181*+Y242F+E259G+N279S+G303R+Y362V+E390A+G475K、 H1*+G7S+R180*+S181*+Y242F+E259G+F261S+N279S+G303R+Y362V+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+F261S+N279S+S286K+G303R+Y362V+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+S286L+G303R+E390A+G475K、 H1*+G7A+G110K+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+W140S+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+W167P+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+H323A+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+W283N+G303R+H320T+H323A+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+H320T+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+P321H+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+M322Y+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+Q360Y+E390A+G475K、 H1*+G7S+T40N+Q98L+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+Q98L+R180*+S181*+Y242F+E259G+F261K+N279S+G303R+Y362V+E390A+G475K、 H1*+G7A+Q98L+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+F261S+N279S+G303R+Y362V+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+S286L+G303R+H320T+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+F261S+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+N194F+L205V+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182D+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+F116N+W140Y+R180*+S181*+T182D+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+F116N+W140Y+R180*+S181*+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+E189P+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+T134D+W140Y+R180*+S181*+E189P+N194F+L205V+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+F116N+W140Y+R180*+S181*+T182D+E189P+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182S+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182S+N194F+L205V+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+F116N+W140Y+R180*+S181*+T182S+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182S+E189P+N194F+L205V+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+F116N+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D123N+W140Y+R180*+S181*+N194F+L205V+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+D123N+W140Y+I176V+R180*+S181*+N194F+L205V+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+G113S+F116W+D123N+W140Y+R180*+S181*+N194F+L205V+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+F116L+D123N+N126D+T131I+W140Y+R180*+S181*+N194F+F202Y+L205V+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+N106D+A109G+W140Y+R180*+S181*+N194F+F202Y+L205V+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+D123N+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+D123N+W140Y+I176V+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+G113S+F116W+D123N+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+F116L+D123N+N126D+T131I+W140Y+R180*+S181*+N194F+F202Y+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+Q98L+G110K+P146S+R180*+S181*+Y242F+E259G+N279S+G303R+A383D+E390A+G475K、 H1*+N3S+G7A+Q98L+G110K+R180*+S181*+Y242F+V252I+E259G+N279S+G303R+E390A+G475K、 H1*+G7H+Q98L+R180*+S181*+Y242F+E259G+N279S+G303R+A383D+E390A+D405G+G475K、 H1*+G7H+Q98L+G110K+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+Q98L+G110K+S132D+Y135E+R180*+S181*+Y242F+E259G+N279S+G303R+T315A+E390A+G475K、 H1*+G7H+G110K+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+A60V+Q98L+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+A383D+E390A+G475K、 H1*+G7A+W140Y+T151A+R180*+S181*+N194F+L205Y+Y242F+K258V+E259V+N279S+G303R+E390A+G475K、 H1*+N3S+G7A+N94D+Q98L+W140Y+R180*+S181*+N194F+L205Y+Y242F+V252I+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+Q98L+G110K+R180*+S181*+Y242F+T256A+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+Q98L+G110K+R180*+S181*+Y242F+E259G+N279S+G303R+A383D+E390A+G475K、 H1*+N3S+G7A+N94D+Q98L+G110K+R180*+S181*+Y242F+V252I+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+Q98L+G110K+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7H+Q98L+G110K+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+T151A+R180*+S181*+N194F+L205Y+Y242F+K258V+E259V+N279S+G303R+E390A+G475K、 H1*+G7A+Q98L+G110K+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+S286L+G303R+Y362V+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+W283N+S286K+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+W283N+G303R+H320T+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+W283N+G303R+H323A+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+S286K+G303R+H320T+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+S286K+G303R+Y362V+E390A+G475K、 H1*+G7S+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+Q98L+R180*+S181*+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+F261K+N279S+G303R+E390A+G475K、 H1*+G7A+R180*+S181*+Y242F+E259G+N279S+G303R+M322G+E359W+E390A+G475K、 H1*+G7A+F116N+W140Y+R180*+S181*+T182D+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+F116N+W140Y+R180*+S181*+T182D+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+F116N+W140Y+R180*+S181*+T182S+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+F116N+W140Y+R180*+S181*+T182D+N194F+L205Y+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+F116N+W140Y+R180*+S181*+A185Q+N194F+L205Y+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+F116N+W140Y+R180*+S181*+T182D+A185Q+N194F+L205Y+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+F116N+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+F116N+W140Y+R180*+S181*+A185Q+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+F116N+W140Y+R180*+S181*+T182D+A185Q+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182D+N194F+L205Y+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+A185Q+N194F+L205Y+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182D+A185Q+N194F+L205Y+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+A185Q+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182D+A185Q+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+E189P+N194F+L205V+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+E189P+N194F+L205V+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182S+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+N194F+L205V+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+N194F+L205V+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+A185Q+N194F+L205V+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+N194F+L205V+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+N194F+L205V+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+A185Q+N194F+L205V+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182S+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+N194F+L205Y+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+N194F+L205Y+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+A185Q+N194F+L205Y+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+A185Q+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+F116N+W140Y+R180*+S181*+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182D+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182D+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+S302T+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+N474F+G475K、 H1*+G7A+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+N422V+G475K、 H1*+G7A+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G476A、 H1*+G7A+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+N394W+G475K、 H1*+G7A+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+N474L、 H1*+G7A+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475E、 H1*+G7A+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+F261P+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+N474F、 H1*+G7A+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+N470A+G475K、 H1*+G7A+W140Y+R180*+S181*+N194F+L205Y+Y242F+T256D+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+N194F+L205Y+Y242L+E259G+N279S+G303R+W346N+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+H420P+G475K、 H1*+G7A+R26Y+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+R37A+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+G475K、 H1*+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+Q360Y+E390A+G475K、 H1*+G7A+R37N+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+G475K、 H1*+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+R414M+G475K、 H1*+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+F261P+N279S+G303R+E390A+G475K、 H1*+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+H323W+E390A+G475K、 H1*+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+P321Y+E390A+G475K、 H1*+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+Q360M+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+Y371S+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+A185Q+N194F+Y242F+S243Q+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+Y242F+S243Q+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+N194F+Y242F+S243Q+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+A185Q+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+E189P+Y242F+S243Q+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+T427L+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+E189P+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+A185Q+E189P+N194F+Y242F+S243Q+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+A185Q+E189P+N194F+L205Y+Y242F+S243Q+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+E189P+L205Y+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+E189P+N194F+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+E189P+N194F+Y242F+S243Q+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+E189P+N194F+L205Y+Y242F+S243Q+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+A185Q+E189P+N194F+L205Y+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+A185Q+E189P+N194F+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+Y242F+S243Q+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+A185Q+Y242F+S243Q+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+Y242F+S243Q+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+N194F+Y242F+S243Q+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+N194F+Y242F+S243Q+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+E189P+Y242F+S243Q+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+Y242F+S243Q+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+E189P+L205Y+Y242F+S243Q+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+A185Q+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+A185Q+N194F+Y242F+S243Q+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+N194F+Y242F+S243Q+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+E189P+N194F+Y242F+S243Q+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+E189P+N194F+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+E189P+N194F+L205Y+Y242F+S243Q+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+A185Q+N194F+Y242F+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+N194F+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+A185Q+E189P+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+Y242F+S243Q+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182D+E189P+L205Y+Y242F+S243Q+E259G+N279S+K280Y+G303R+E390A+I409L+G475K、 H1*+G7A+A41D+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+T40G+W140Y+R180*+S181*+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 W140Y+L173F+R180*+S181*+N194F+L205Y+N225S+Y242F+E259G+N279S+G303R+H323A、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+Y362V+E390A+H420A+N474L、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+N474L、 H1*+N3D+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394D+N474F、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394W+H420P+N474L、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G476A、 H1*+N3D+G7A+K93R+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+N474L、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390D+H420P+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394W+H420A+G475K、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394D+H420A+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420P+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+N474L、 N3G+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+N474L、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394W+H420A+G475K、 H1*+N3D+G7A+R37N+D118T+Q138V+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394W+H420A+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420P+G475K、 H1*+G7A+D118T+Q138V+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+H323W+E390A+N474L、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+F261P+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+N474L、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+P321H+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+P321H+E390A+N474L、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394D+R414M+G475K、 H1*+G7A+T40G+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+T224F+Y242F+E259G+N279S+K280Y+G303R+N394W+H420A+G475K、 H1*+G7A+D118T+Y135E+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+W346N+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+P321H+Q360Y+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260W+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+T224Y+Y242F+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260W+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+T224Y+Y242F+E259G+N279S+K280Y+G303R+E390A+N474L、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+S132L+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205V+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205N+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Q+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+A185Q+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+N194F+L205Y+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+N194F+L205N+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185D+E189P+N194F+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185D+E189P+Y242F+S243Q+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180Q+S181*+T182*+A185Q+E189P+Y242F+S243Q+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+L205Y+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185D+E189P+L205Y+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180Q+S181*+T182*+A185Q+E189P+N194F+L205Y+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+T40G+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+T40G+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394D+N422V+G475K、 H1*+G7A+Q98L+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394D+H420A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+Y362V+E390A+G475K、 H1*+G7A+D118T+Y135E+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+F261P+N279S+K280Y+G303R+Y362V+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+T256D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+Y135E+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+T224F+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+T256D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+T256D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+F261P+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+T256D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+Y371N+E390A+G475K、 H1*+G7A+D118T+Y135E+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+P321Y+E390A+G475K、 H1*+G7A+D118T+S132D+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+P321Y+E390A+G475K、 H1*+N3G+G7A+D118T+Q138V+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+T256D+N279S+K280Y+G303R+E390A+N470M+G475K、 H1*+G7A+D118T+S132D+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+Y362V+E390A+N474L、 H1*+G7A+D118T+Q138V+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+Y362V+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+N474L、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+T224Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+N474F、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+H420A+G475K、 H1*+G7A+D118T+Y135E+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394W+H420A+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394D+H420A+G475K、 H1*+G7A+D118T+Q138V+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420Q+G475K、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420P+G476H、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420P+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+N470M+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+H420P+G476A、 H1*+G7A+R37N+D118T+S132D+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+N470M+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+H323W+E390A+H420A+N474L、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394W+H420P+G475K、 H1*+G7A+D118T+Y135E+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+R414M+G476H、 H1*+N3D+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G475K、 H1*+N3D+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G475K、 H1*+N3D+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+N3G+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394W+H420A+G475K、 H1*+N3D+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G475K、 H1*+N3D+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+G361M+H420A+G475K、 H1*+N3D+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+N394W+G475K、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+T224F+Y242F+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+N394D+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+N394D+G475K、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+N394D+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+H420A+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+T224Y+Y242F+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394W+H420A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394W+H420A+N474L、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+N194F+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+N194F+L205Q+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180Q+S181*+T182*+A185Q+E189P+N194F+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+Y242F+S243Q+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+N3G+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+N394W+H420A+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G475K、 H1*+N3D+G7A+D118T+S132D+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+N470M+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+N394W+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260W+N279S+K280Y+G303R+E390A+N470M+G475K、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+N394D+G475K、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260W+N279S+K280Y+G303R+E390A+H420A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+N470M+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+N394W+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+T224Y+Y242F+E259G+N279S+K280Y+G303R+N394W+G475K、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+N394W+H420A+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+N470M+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+T256D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+F261P+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+N394D+H420A+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+N394D+G475K、 H1*+G7A+R37N+D118T+W140Y+N174K+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+N394W+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+T224Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+T224Y+Y242F+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+H420A+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+T224Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+T224Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+T224Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+N3G+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+T256D+N279S+K280Y+G303R+E390A+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+T256D+N279S+K280Y+G303R+E390A+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+T256D+N279S+K280Y+G303R+E390A+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+T256D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+T224F+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G476H、 H1*+G7A+Q98L+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+T256D+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+N394D+G475K、 N3G+G7A+R37V+D118T+W140M+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+E390A+H420A+G475K、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+H323W+N394D+G475K、 H1*+G7A+R37A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+P321Y+L354V+N394D+G475K、 H1*+G7A+R37N+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+H323W+Q360M+E390A+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+H323W+E390A+G475K、 H1*+N3D+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+L205Y+Y242F+E259G+N279S+K280Y+G303R+H323W+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185D+E189P+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180Q+S181*+T182*+A185Q+E189P+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+Y242F+E259G+M260W+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182D+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+S132L+W140Y+R180*+S181*+T182D+E189P+N194F+L205Y+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+N194F+L205Y+T224Y+Y242F+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+N194F+Y242F+E259G+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+N194F+L205Y+Y242F+E259G+M260W+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+T224Y+Y242F+F266Y+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+S132L+W140Y+R180*+S181*+T182S+E189P+N194F+Y242F+E259G+M260D+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+Y242F+E259G+M260W+F266Y+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+N194F+T224Y+Y242F+F266Y+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+N194F+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+S132L+W140Y+R180*+S181*+T182S+A185Q+E189P+N194F+L205Y+Y242F+E259G+M260D+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+N194F+Y242F+E259G+M260W+F266Y+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+W140Y+R180*+S181*+E189P+N194F+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182D+E189P+N194F+Y242F+E259G+M260W+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+N194F+Y242F+E259G+M260W+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+T224Y+Y242F+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182D+E189P+N194F+T224Y+Y242F+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+N194F+T224Y+Y242F+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182D+E189P+N194F+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182D+A185Q+E189P+N194F+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182D+A185Q+Y242F+S243Q+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182D+N194F+Y242F+S243Q+E259G+F266Y+N279S+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+N196I+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185D+E189P+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+H420Y+G475K、 H1*+G7A+D118T+W140Y+R180*+S181*+A185Q+E189P+N194F+L205Y+Y242F+E259G+F266Y+N279S+K280Y+G303R+E390A+G475K、 H1*+G7A+W140Y+R180*+S181*+E189P+N194F+Y242F+E259G+M260W+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182D+E189P+N194F+T224Y+Y242F+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182S+E189P+N194F+T224Y+Y242F+N279S+K280Y+G303R+E390A+G476A、 H1*+G7A+D118T+W140Y+R180*+S181*+T182D+A185Q+E189P+N194F+T224Y+Y242F+N279S+K280Y+G303R+E390A+G476A, H1*+G7A+S132L+W140Y+R180*+S181*+E189P+N194F+Y242F+E259G+M260D+N279S+K280Y+G303R+E390A+G475K, H1*+G7A+D118T+W140Y+R180*+S181*+T182S+A185Q+E189P+N194F+Y242F+E259G+M260W+N279S+K280Y+G303R+E390A+G476A, H1*+G7A+D118T+W140Y+R180*+S181*+T182D+E189P+N194F+L205Y+T224Y+Y242F+F266Y+N279S+K280Y+G303R+E390A+G476A, and one or more substitutions or deletions selected from the group consisting of:

6. Use of an α-amylase variant according to any one of claims 1 to 5 in cleaning processes such as hard surface cleaning and industrial cleaning, including laundry or dishwashing.

7. A composition comprising the mutant of any one of claims 1 to 5.

8. 8. The composition of claim 7, wherein the composition is selected from the group comprising liquid detergents, solid detergents, gel detergents, powder detergents, tubs, pods, pouches and granular detergents.

9. 9. The composition of claim 7 or 8, which is a liquid laundry or liquid dishwashing composition, such as an automatic dishwashing (ADW) liquid detergent composition, or a solid laundry composition, such as a soap bar, or a powder dishwashing composition, such as an ADW unit dose detergent composition.

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