Corynebacterium mutant microorganisms that produce L-glutamic acid and a method for producing L-glutamic acid using the same

Abstract

The present invention relates to a Corynebacterium mutant microorganism that produces L-glutamic acid and a method for producing L-glutamic acid using the same. More specifically, the present invention relates to a novel mutant of an FAD-dependent oxidoreductase involved in the biosynthetic pathway of L-glutamic acid, a polynucleotide, a transformant, and a method for producing L-glutamic acid using the same. The FAD-dependent oxidoreductase mutant according to the present invention has a change in enzyme activity due to substitution of one or more amino acids in the amino acid sequence constituting the FAD-dependent oxidoreductase, and a recombinant microorganism containing the same can efficiently produce L-glutamic acid.

Description

[Technical Field]

[0001] The present invention relates to a mutant Corynebacterium microorganism that produces L-glutamic acid and a method for producing L-glutamic acid using the same, and more specifically relates to a novel mutant of FAD-dependent oxidoreductase involved in the biosynthetic pathway of L-glutamic acid, a polynucleotide, a transformant, and a method for producing L-glutamic acid using the same. [Background Art]

[0002] L-glutamic acid is a representative amino acid produced by microbial fermentation. Monosodium L-glutamate (MSG) balances and harmonizes the overall flavor of food, enhances the preference of foods such as meat, fish, chicken, vegetables, sauces, soups and seasonings, can improve the taste of low-salt foods with a 30% salt reduction, and is widely used as a seasoning for household use and processed food production.

[0003] A brief look at the fermentation pathway of L-glutamic acid reveals that glucose is primarily metabolized via the glycolytic pathway, but some is also metabolized via the pentose phosphate pathway into two molecules of pyruvic acid. One of these molecules fixes CO2 to become oxaloacetic acid, while the other combines with acetyl-CoA to form citric acid. Furthermore, oxaloacetic acid and citric acid enter the citric acid cycle (TCA cycle) to become alpha-ketoglutaric acid. Here, the oxidative metabolic pathway that oxidizes alpha-ketoglutaric acid to succinic acid is absent, and because isocitrate dehydrogenase and glutamate dehydrogenase are closely involved, the reductive amino acid conversion reaction of alpha-ketoglutaric acid proceeds efficiently to produce L-glutamic acid.

[0004] L-glutamic acid can be produced using wild-type strains obtained in nature or mutant strains modified to enhance their glutamic acid production capacity. Recently, in order to improve the efficiency of L-glutamic acid production, genetic engineering techniques have been applied to microorganisms such as Escherichia coli and Corynebacterium, which are widely used in the production of useful substances such as amino acids and nucleic acids, to develop diverse recombinant or mutant strains with superior L-glutamic acid production capabilities and methods for producing L-glutamic acid using these. In particular, there have been attempts to increase L-glutamic acid production by targeting genes such as enzymes, transcription factors, and transport proteins involved in the L-glutamic acid biosynthesis pathway, or by inducing mutations in promoters that regulate their expression. However, since there are dozens of types of proteins, including enzymes, transcription factors, and transport proteins, that are directly or indirectly related to L-glutamic acid production, much research is still needed to determine whether changes in the activity of such proteins increase L-glutamic acid production capacity. [Prior art documents] [Patent Documents]

[0005] [Patent Document 1] U.S. Registered Patent No. 6,852,516 [Patent Document 2] U.S. Registered Patent No. 6,962,805 [Overview of the project] [Problems that the invention aims to solve]

[0006] The present invention aims to provide novel FAD-dependent oxidoreductase variants.

[0007] Furthermore, the present invention relates to the above variants code The objective is to provide polynucleotides.

[0008] Furthermore, the present invention aims to provide a transformant comprising the aforementioned mutant or polynucleotide.

[0009] Furthermore, the present invention aims to provide a method for producing L-glutamic acid using the aforementioned transformant. [Means for solving the problem]

[0010] One aspect of the present invention provides a FAD-dependent oxidoreductase mutant comprising the amino acid sequence of SEQ ID NO: 2, wherein the 200th alanine in the amino acid sequence of SEQ ID NO: 4 is replaced with threonine.

[0011] The "FAD-dependent oxidoreductase" used in this invention is an enzyme that catalyzes redox reactions to supply energy necessary for living organisms, and it oxidizes one compound while reducing other compounds using FAD. The FAD-dependent oxidoreductase is codeThis may be the gene or a sequence having substantial identity therewith. Here, "substantial identity" means that when each gene sequence, i.e., the base sequence or nucleotide sequence, is aligned with any other nucleotide sequence in a manner that maximizes correspondence and analyzed, the other nucleotide sequence has a sequence homology of 70% or more, 80% or more, 90% or more, or 98% or more with each nucleotide sequence.

[0012] The FAD-dependent oxidoreductase in this invention contains the amino acid sequence of SEQ ID NO: 4.

[0013] According to one specific example of the present invention, the amino acid sequence of Sequence ID No. 4 may be derived from a wild-type Corynebacterium microorganism.

[0014] More specifically, the Corynebacterium microorganism may be Corynebacterium glutamicum.

[0015] The term "mutant" as used in this invention refers to a polypeptide in which one or more amino acids in the N-terminus, C-terminus, and / or internal part of the amino acid sequence of a specific gene are conservatively substituted, deleted, modified, or added, resulting in a sequence that differs from the amino acid sequence of the mutant before the mutation, but maintains its functions or properties. Here, "conservative substitution" means substituting one amino acid with another amino acid that has similar structural and / or chemical properties, and has little to no effect on the activity of the protein or polypeptide. The aforementioned amino acids were selected from alanine (Ala), isoleucine (Ile), valine (Val), leucine (Leu), methionine (Met), asparagine (Asn), cysteine ​​(Cys), glutamine (Gln), serine (Ser), threonine (Thr), phenylalanine (Phe), tryptophan (Trp), tyrosine (Tyr), aspartic acid (Asp), glutamic acid (Glu), arginine (Arg), histidine (His), lysine (Lys), glycine (Gly), and proline (Pro).

[0016] Furthermore, mutants may include those in which one or more parts, such as the N-terminal leader sequence or transmembrane domain, are removed, or in which a portion of the N- and / or C-terminus of a mature protein is removed.

[0017] Such mutants may have increased (enhanced), unchanged, or decreased (weakened) capabilities compared to the pre-mutation protein or polypeptide. Here, "increased or enhanced" means that the activity of the protein itself has increased compared to the pre-mutation protein. codeWhen the overall level of protein activity in the cell is higher than that of a wild-type strain or a strain expressing the protein before mutation due to increased expression or increased translation of the gene, and includes combinations thereof. The term "reduced or weakened" includes when the activity of the protein itself is reduced compared to that of the protein before mutation, and when the overall level of protein activity in the cell is lower than that of a wild-type strain or a strain expressing the protein before mutation due to inhibition of expression or inhibition of translation of the gene that encodes the protein code , and includes combinations of the above cases. In the present invention, the term "variant" may be used interchangeably with mutant, variant form, variant polypeptide, mutated protein, mutation, etc.

[0018] The variant of the present invention is a FAD-dependent oxidoreductase in which alanine, which is the 200th amino acid in the amino acid sequence of SEQ ID NO: 4, is substituted with threonine, and may consist of the amino acid sequence of SEQ ID NO: 2.

[0019] Another aspect of the present invention provides a polynucleotide encoding the FAD-dependent oxidoreductase variant code .

[0020] As used in the present invention, a "polynucleotide" is a polymer of nucleotides in which nucleotide monomers are covalently linked in a long chain, and is a DNA or RNA chain of at least a certain length; more specifically, it refers to a polynucleotide fragment encoding the variant code .

[0021] The polynucleotide may comprise a nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 2 code .

[0022] According to one specific embodiment of the present invention, the polynucleotide may comprise the nucleotide sequence represented by SEQ ID NO: 1.

[0023] Another aspect of the present invention relates to the FAD-dependent oxidoreductase variant codeProvided is a vector comprising the polynucleotide.

[0024] In another aspect, the present invention provides a transformant comprising the FAD-dependent oxidoreductase variant or the polynucleotide.

[0025] As used in the present invention, the "vector" refers to all types of nucleic acid sequence delivery constructs used as a means for delivering and expressing a target gene in a host cell. Unless otherwise specified, the vector may refer to a vector in which the carried nucleic acid sequence is inserted into a host cell genome for expression and / or expressed independently. Such a vector comprises essential regulatory elements operably linked to enable expression of the gene insert. The term "operably linked" means that a target gene and its regulatory sequences are functionally linked to each other in a manner that allows gene expression, and the term "regulatory element" includes a promoter for transcription, any operator sequence for regulating transcription, and a suitable mRNA ribosome binding site code and sequences that regulate the termination of transcription and translation.

[0026] The vector used in the present invention is not particularly limited as long as it is replicable in a host cell, and any vector known in the art can be used. Examples of the vector include plasmids in natural or recombinant state, cosmids, viruses and bacteriophages. For example, examples of phage vectors or cosmid vectors include, but are not limited to, pWE15, M13, λMBL3, λMBL4, λIXII, λASHII, λAPII, λt10, λt11, Charon4A and Charon21A; examples of plasmid vectors include pBR series, pUC series, pBluescriptII series, pGEM series, pTZ series, pCL series and pET series, but are not limited thereto.

[0027] The aforementioned vectors can typically be constructed as vectors for cloning or vectors for expression. For expression, standard vectors used in the art to express foreign genes or proteins in plants, animals, or microorganisms can be used, and can be constructed by a variety of methods known in the art.

[0028] The "recombinant vector" used in this invention may, after being transformed into a suitable host cell, be able to replicate independently of the host cell's genome or be stitched into the genome itself. In this case, the "suitable host cell" is one in which the vector can replicate and may include an origin of replication, which is a specific base sequence from which replication is initiated. For example, if the vector used is an expression vector and the host is a prokaryotic cell, it typically includes a strong promoter that can advance transcription (e.g., pLλ promoter, CMV promoter, trp promoter, lac promoter, tac promoter, T7 promoter), a ribosome binding site for initiating sequencing, and a transcription / sequence termination sequence. If the host is a eukaryotic cell, the origins of replication that act in eukaryotic cells and are included in the vector include, but are not limited to, the f1 origin of replication, SV40 origin of replication, pMB1 origin of replication, adeno origin of replication, AAV origin of replication, and BBV origin of replication. Furthermore, promoters derived from the genome of mammalian cells (e.g., metallothione promoter) or promoters derived from mammalian viruses (e.g., late adenovirus promoter, vaccinia virus 7.5K promoter, SV40 promoter, cytomegalovirus promoter, HSV tk promoter) may be used, and generally have a polyadenylated sequence as the transcription termination sequence.

[0029] The recombinant vector may contain a selection marker, which is used to select transformants (host cells) transformed by the vector. Since only cells expressing the selection marker can survive in a culture medium treated with the selection marker, the transformed cells can be selected. Typical examples of the selection marker include, but are not limited to, kanamycin, streptomycin, and chloramphenicol.

[0030] A transformant can be created by inserting a recombinant vector into a host cell, and the transformant may be obtained by introducing the recombinant vector into a suitable host cell. The host cell is a cell that can stably and continuously clone or express the expression vector, and any host cell known in the art can be used.

[0031] When transforming prokaryotic cells to produce recombinant microorganisms, the host cells may include, but are not limited to, a variety of Enterobacteria and bacterial strains such as E. coli strains like E. coli JM109, E. coli BL21, E. coli RR1, E. coli LE392, E. coli B, E. coli X 1776, E. coli W3110, and E. coli XL1-Blue, Bacillus strains like Bacillus subtilis and Bacillus thuringiensis, Corynebacterium strains, Salmonella tiphyllum, Serratia marcescens, and Pseudomonas species.

[0032] When transforming eukaryotic cells to produce recombinant microorganisms, the host cells may include, but are not limited to, yeast (e.g., Saccharomyces cerevisiae), insect cells, plant cells, and animal cells, such as Sp2 / 0, CHO K1, CHO DG44, PER.C6, W138, BHK, COS7, 293, HepG2, Huh7, 3T3, RIN, and MDCK cell lines.

[0033] In this invention, "transformation" refers to the phenomenon of artificially inducing genetic changes by introducing external DNA into host cells, while "transformatted organism" refers to a host cell into which external DNA has been introduced and which maintains stable expression of the target gene.

[0034] The transformation can be performed by selecting a suitable vector introduction technique for the host cell to express the target gene or a recombinant vector containing it within the host cell. For example, vector introduction may be performed by electroporation, heat shock, calcium phosphate (CaPO4) precipitation, calcium chloride (CaCl2) precipitation, microinjection, polyethylene glycol (PEG) method, DEAE-dextran method, cation liposome method, lithium acetate-DMSO method, or a combination thereof, but is not limited to these. The transformed gene can be expressed within the host cell, and is not limited to those located inside or outside the host cell's chromosome.

[0035] The transformants include cells that have been transfected, transformed, or infected with the recombinant vector according to the present invention in vivo or in vitro, and can be used as synonymous with recombinant host cells, recombinant cells, or recombinant microorganisms.

[0036] According to one specific example of the present invention, the transformed organism may be a microorganism of the genus Corynebacterium.

[0037] More specifically, the Corynebacterium genus microorganisms mentioned above include Corynebacterium glutamicum, Corynebacterium crudilactis, Corynebacterium deserti, Corynebacterium callunae, Corynebacterium suranareeae, Corynebacterium lubricantis, Corynebacterium doosanense, Corynebacterium efficiens, and Corynebacterium uterechii. Corynebacterium uterequi), Corynebacterium stationis, Corynebacterium pacaense, Corynebacterium singulare, Corynebacterium humireducens, Corynebacterium marinum, Corynebacterium halotolerans, Corynebacterium spheniscorum, Corynebacterium freiburgense, Corynebacterium striatum (Corynebacterium Corynebacterium striatum), Corynebacterium canis, Corynebacterium ammoniagenes, Corynebacterium renale, Corynebacterium portisoliExamples include, but are not limited to, Corynebacterium pollutisoli, Corynebacterium imitans, Corynebacterium caspium, Corynebacterium testudinoris, Corynebacterium pseudopelargi, or Corynebacterium flavescens.

[0038] The transformant in this invention is the aforementioned FAD-dependent oxidoreductase mutant or the transformed organism. code This may include, but is not limited to, a strain containing a polynucleotide or a vector containing the same, a strain expressing the FAD-dependent oxidoreductase mutant or polynucleotide, or a strain having activity against the FAD-dependent oxidoreductase mutant.

[0039] According to one specific example of the present invention, the transformant may have the ability to produce L-glutamic acid.

[0040] The transformed organism may naturally possess the ability to produce L-glutamate, or it may have been artificially given the ability to produce L-glutamate.

[0041] According to one specific example of the present invention, the transformant may have altered FAD-dependent oxidoreductase activity, resulting in improved L-glutamic acid production capacity.

[0042] In this invention, "improved productivity" means an increase in L-glutamic acid production compared to the parent strain. The parent strain refers to the wild type or mutant strain that is the target of mutation, and includes subjects that are directly targeted for mutation or transformed with recombinant vectors, etc. In this invention, the parent strain may be a wild-type Corynebacterium microorganism or a Corynebacterium microorganism that has mutated from the wild type.

[0043] The transformants according to the present invention exhibit increased L-glutamic acid production capacity compared to the parent strain, due to the introduction of a FAD-dependent oxidoreductase mutant, which alters the activity of the FAD-dependent oxidoreductase. More specifically, the transformants may, but are not limited to, those whose L-glutamic acid production is increased by at least 1%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100% compared to the parent strain, or by 1.1 times, 1.5 times, 2 times, 2.5 times, 3 times, 3.5 times, 4 times, 4.5 times, 5 times, 5.5 times, 6 times, 6.5 times, 7 times, 7.5 times, 8 times, 8.5 times, 9 times, 9.5 times, or 10 times. As an example, the transformant containing the FAD-dependent oxidoreductase mutant may have a L-glutamic acid production that is 5% or more, specifically 5-50% (preferably 7-30%) higher than that of the parent strain.

[0044] Another aspect of the present invention provides a method for producing L-glutamic acid, comprising the steps of culturing the transformant in a culture medium and recovering L-glutamic acid from the transformant or the culture medium in which the transformant was cultured.

[0045] The culture may be carried out using appropriate culture media and conditions known in the art, or a typical technician can easily adjust and use the culture media and conditions. Specifically, the culture media may be, but is not limited to, a liquid culture. The culture method may include, but is not limited to, batch culture, continuous culture, fed-batch culture, or a combination thereof.

[0046] According to one specific example of the present invention, the culture medium must meet the requirements of a specific bacterial strain in an appropriate manner and can be modified as appropriate by an ordinary technician. Culture media for microorganisms of the genus Corynebacterium can be found in, but are not limited to, known literature (Manual of Methods for General Bacteriology. American Society for Bacteriology. Washington DC, USA, 1981).

[0047] According to one specific example of the present invention, the culture medium may contain a variety of carbon sources, nitrogen sources, and trace element components. Usable carbon sources include sugars and carbohydrates such as glucose, sucrose, lactose, fructose, maltose, starch, and cellulose; oils and fats such as soybean oil, sunflower oil, castor oil, and coconut oil; fatty acids such as palmitic acid, stearic acid, and linoleic acid; alcohols such as glycerol and ethanol; and organic acids such as acetic acid. These substances may be used individually or in mixtures, but are not limited thereto. Usable nitrogen sources may include peptone, yeast extract, meat juice, malt extract, corn maceration, soybean meal, and urea, or inorganic compounds such as ammonium sulfate, ammonium chloride, ammonium phosphate, ammonium carbonate, and ammonium nitrate. Nitrogen sources may also be used individually or in mixtures, but are not limited thereto. Usable phosphorus sources may include, but are not limited thereto, potassium dihydrogen phosphate or dipotassium hydrogen phosphate or corresponding sodium-containing salts. Furthermore, the culture medium may, but is not limited to, contain metal salts such as magnesium sulfate or iron sulfate necessary for growth. Other essential growth substances such as amino acids and vitamins may also be included. In addition, suitable precursors can be used in the culture medium. The medium or individual components may, but are not limited to, be added to the culture medium in a batch or continuous manner in a manner appropriate to the culture process.

[0048] According to one specific example of the present invention, the pH of the microbial culture medium can be adjusted by adding compounds such as ammonium hydroxide, potassium hydroxide, ammonia, phosphoric acid, and sulfuric acid to the culture medium in an appropriate manner during cultivation. Furthermore, the formation of bubbles can be suppressed during cultivation using an antifoaming agent such as fatty acid polyglycol ester. Additionally, oxygen or an oxygen-containing gas (e.g., air) can be injected into the culture medium to maintain an aerobic state. The temperature of the culture medium is typically 20-45°C, for example, 25-40°C. The cultivation period can be continued until the useful substance is obtained in the desired yield, for example, 10-160 hours.

[0049] According to one specific example of the present invention, the step of recovering L-glutamic acid from the cultured transformant or the culture medium in which the transformant was cultured can be performed by collecting or recovering the L-glutamic acid produced from the culture medium using a suitable method known in the art, depending on the culture method. For example, methods such as centrifugation, filtration, extraction, spraying, drying, evaporation, precipitation, crystallization, electrophoresis, fractional dissolution (e.g., ammonium sulfate precipitation), and chromatography (e.g., ion exchange, affinity, hydrophobicity, and size exclusion) can be used, but are not limited thereto.

[0050] According to one specific example of the present invention, the step of recovering L-glutamic acid can be performed by removing biomass by slow centrifugation of the culture medium and separating the resulting supernatant by ion exchange chromatography.

[0051] According to one specific example of the present invention, the step of recovering L-glutamic acid may include a step of purifying L-glutamic acid. [Effects of the Invention]

[0052] The FAD-dependent oxidoreductase mutant according to the present invention has altered enzyme activity due to the substitution of one or more amino acids in the amino acid sequence constituting the FAD-dependent oxidoreductase, and recombinant microorganisms containing this mutant can efficiently produce L-glutamic acid. [Brief explanation of the drawing]

[0053] [Figure 1] Figure 1 shows the structure of plasmid pK19msb according to one embodiment of the present invention. [Modes for carrying out the invention]

[0054] The present invention will be described in more detail below. However, such description is provided only as an example for the purpose of understanding the present invention, and the scope of the present invention is not limited by such exemplary description.

[0055] Example 1. Vector preparation for FAD-dependent oxidoreductase mutant expression. We created a vector expressing a mutant of FAD-dependent oxidoreductase in which the alanine (A) at position 200 in the amino acid sequence (SEQ ID NO: 4) is replaced with threonine (T).

[0056] Using the gDNA of wild-type Corynebacterium glutamicum ATCC13869 as a template, PCR was performed using primer pairs 1 and 2, and primer pairs 3 and 4. Subsequently, a mixture of the two PCR products was used as a template, and overlapping PCR was performed again using primers 1 and 4 to obtain fragments. Here, Takara PrimeSTAR Max DNA polymerase was used as the polymerase, and the PCR amplification conditions were denaturation at 95°C for 5 minutes, followed by 30 repetitions of 95°C for 30 seconds, 58°C for 30 seconds, and 72°C for 1 minute 30 seconds, followed by a reaction at 72°C for 5 minutes. The pK19msb vector was treated with smaI and ligated with the obtained PCR product (fragment), and the plasmid obtained was named pK_fprA(A200T).

[0057] The primer sequences used to prepare the vector are shown in Table 1 below.

[0058] [Table 1]

[0059] Example 2. Creation of mutant strains into which FAD-dependent oxidoreductase mutants have been introduced. Corynebacterium glutamicum U3 (KCCM13218P) was used as the parent strain to introduce FAD-dependent oxidoreductase mutants, and a modified electrocompetent cell production method based on van der Rest's method was used for transforming the U3 strain.

[0060] First, the U3 strain was cultured in 10 ml of 2YT medium (containing 16 g / l tryptone, 10 g / l yeast extract, and 5 g / l sodium chloride) supplemented with 2% glucose to prepare a seed culture. Subsequently, 100 ml of 2YT medium (without glucose) was inoculated with 1 mg / ml isonicotinic acid hydrazine and 2.5% glycine to achieve an OD610 value of 0.3. The culture was then incubated at 18°C ​​and 180 rpm for 12-16 hours until the OD610 value reached 1.2-1.4. After leaving the culture on ice for 30 minutes, it was centrifuged at 4°C and 4000 rpm for 15 minutes. The supernatant was then discarded, and the precipitated U3 strain was washed four times with 10% glycerol solution. Finally, it was resuspended in 0.5 ml of 10% glycerol solution to prepare competent cells. Electroporation was performed using an electroporator from Bio-Rad. After adding the prepared competent cells and pK_fprA(A200T) vector to an electroporation cuvette (0.2 mm), an electric shock was applied under conditions of 2.5 kV, 200 Ω, and 12.5 μF. Immediately after the electric shock, 1 ml of regeneration medium (containing Brain Heart infusion 18.5 g / l and sorbitol 0.5 M) was added, and the cells were heat-treated at 46 °C for 6 minutes. After cooling to room temperature, the mixture was transferred to a 15 ml capped tube and incubated at 30°C for 2 hours. The mixture was then streaked onto a selection medium (containing 5 g / l tryptone, 5 g / l NaCl, 2.5 g / l yeast extract, 18.5 g / l Brain Heart infusion powder, 15 g / l agar, 91 g / l sorbitol, and 20 μg / l kanamycin). Colonies were incubated at 30°C for 72 hours to induce secondary recombination in BHI medium. -5 ~10 -7 The sample was diluted to [specific concentration] and spread onto antibiotic-free 2YT agar plates (containing 10% sucrose). Strains that showed no resistance to kanamycin and grew well in a medium containing 10% sucrose were selected and named FPR1.

[0061] Experimental Example 1. Evaluation of L-glutamate production capacity of mutant strains introduced with FAD-dependent oxidoreductase mutants. We compared the L-glutamate production capacity of the parent strain U3 and the mutant strain FPR1, which was introduced with a FAD-dependent oxidoreductase mutant.

[0062] Each bacterial strain (parent strain or mutant strain) was inoculated at a volume of 1% into a 100 mL flask containing 10 mL of glutamic acid production medium (as shown in Table 2 below), and the culture was incubated with shaking at 30°C and 200 rpm for 48 hours. After incubation, the concentration of L-glutamic acid in the medium was measured using HPLC (Agilent), and the results are shown in Table 3 below.

[0063] [Table 2]

[0064] [Table 3]

[0065] As shown in Table 3 above, mutant strains into which FAD-dependent oxidoreductase mutants were introduced showed an approximately 12.8% increase in L-glutamic acid production compared to the parent strain. These results suggest that the FAD-dependent oxidoreductase mutation increases the flow of carbon sources in the glutamic acid production pathway, thereby increasing L-glutamic acid productivity.

[0066] The present invention has been described above, focusing on its preferred embodiments. Those with ordinary skill in the art to which the present invention pertains will understand that the present invention can be realized in modified forms without departing from its essential characteristics. Therefore, the disclosed embodiments should be considered in an explanatory rather than restrictive manner. The scope of the present invention is shown in the claims, not in the above description, and all differences within an equivalent scope should be interpreted as being included in the present invention.

[0067] [Accession Number] Depository name: Korea Center for Microbial Conservation (KCCM) Accession number: KCCM13218P Date of acceptance: 20220629

Claims

1. A transformant having the ability to produce L-glutamic acid and comprising a FAD-dependent oxidoreductase mutant having the amino acid sequence of SEQ ID NO: 2, or a polynucleotide encoding the mutant.

2. The transformant according to claim 1, wherein the microorganism is of the genus Corynebacterium.

3. The step of culturing the transformant according to claim 1 in a culture medium, A method for producing L-glutamic acid, comprising the step of recovering L-glutamic acid from the transformant or the culture medium in which the transformant was cultured.

4. A composition for producing L-glutamic acid, comprising the transformant described in Claim 1.

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