Lipases, compositions, methods, and uses thereof

Abstract

The present invention relates to a wild-type, non-genetically modified, microbial lipolytic enzyme that has a higher specificity for medium- to long-chain fatty acids, particularly short-chain fatty acids, compared to long-chain fatty acids. The present invention further relates to a process for preparing a food product and the use of the wild-type, non-genetically modified, microbial lipolytic enzyme.

Description

[Technical Field]

[0001] The present invention relates to wild-type, non-genetically modified, microbial lipolytic enzymes, nucleic acids encoding said lipolytic enzymes, vectors, host cells and / or compositions comprising said lipolytic enzymes, preferably having a high specificity for short-chain fatty acids compared to medium- to long-chain fatty acids when compared to known microbial lipolytic enzymes. The present invention also relates to processes for preparing food products such as dairy products, in which said lipolytic enzymes are used. The present invention is suitable for use in the food industry, preferably the dairy industry, more preferably the cheese industry, for example, to improve the flavor of ripened cheese or shorten the ripening period. [Background technology]

[0002] background Lipolytic enzymes, also called lipases (EC 3.1.1.3), are a class of hydrolases that hydrolyze the ester bonds of lipid triglycerides to produce free fatty acids. This class of enzymes is involved, for example, in the lipolysis of milk fat and / or other fats with the subsequent release of short (C4- to C6-fatty acids), medium- and / or long (C8- or longer) chain fatty acids.

[0003] Each lipolytic enzyme has its own profile of free fatty acid release from milk fat and / or other lipids. This profile can range from short (C4- to C6-fatty acids), medium- and / or long-chain fatty acids (C8 or longer). The resulting mixture of short (C4- to C6-fatty acids), medium- and / or long-chain fatty acids (C8 or longer) depends, at least, on the lipolytic enzyme and the lipolytic substrate. The resulting mixture of short (C4- to C6-fatty acids), medium- and / or long-chain fatty acids (C8 or longer) contributes to modifying the flavor of a given end product. For example, a mixture with a higher amount or concentration of short-chain fatty acids, such as butyric acid (C4-fatty acid), compared to the amount or concentration of medium- and / or long-chain fatty acids leads to the development of a desirable rancid cheese flavor, especially during ripening, and has been used to flavor dairy products, e.g., various cheese types. In contrast, mixtures having a greater amount or concentration of medium and / or long chain fatty acids compared to the amount or concentration of short chain fatty acids, especially mixtures having a greater amount or concentration of long chain fatty acids (C 16 - or more) will result in an unpleasant odor.

[0004] Lipolytic enzymes or lipases can be of animal or non-animal origin. Lipases of animal origin (kid, calf, or lamb) preferentially cleave short-chain fatty acids (C4- to C6-fatty acids) rather than long-chain fatty acids from milk fat and / or other fats. These short-chain fatty acids, especially C4-fatty acids, are responsible for the formation of a pleasant rancid flavor rather than the soapy flavor produced by medium- to long-chain fatty acids. To a lesser extent, they also release medium-chain fatty acids, which add an animal-specific flavor to cheese. Nevertheless, lipases from animal origin cannot be used in vegetarian and / or kosher products (which are highly desired by the market). On the other hand, lipases from non-animal origin, such as microbial origin, fully meet vegetarian and / or kosher requirements. However, microbial lipases are known to be nonspecific lipases that primarily release medium- to long-chain fatty acids (C8 and above) from milk fat and / or other lipids, resulting in a very unpleasant soapy flavor in cheese.

[0005] Therefore, in the food industry, and in particular in the cheese industry, there is a need to search for new sources of wild-type, non-genetically modified, microbial lipases that exhibit a higher specificity for short-chain fatty acids, such as C4- and / or C6-fatty acids, compared to medium- or long-chain fatty acids, and / or a higher specificity for short-chain fatty acids, such as C4- and / or C6-fatty acids, compared to long-chain fatty acids, and also for C8- or C9-fatty acids. 10 There is an impetus to search for new sources of microbial wild-type, non-genetically modified, lipases that display specificity for medium-chain fatty acids such as -fatty acids.

[0006] Several attempts to replace animal lipases are known in the prior art. These attempts include the use of microbial lipases, recombinant lipases, and / or genetically engineered lipases. However, these attempts have not provided wild-type microbial lipases with higher specificity for short-chain fatty acids, preferably C4-fatty acids, and the desired flavor in food products such as cheese.

[0007] Microbial lipases can be obtained, for example, from Aspergillus, Rhizopus, and Mucor. The effect of these fungal lipases on lipid rancidity is disclosed for Aspergillus by Iwai et al. in J. Gen. Appl. Microbiol. 10, 13-22 (1964), US 4 726 954 or US 4 636 468, US 4 726 954 or US 4 636 468; for Rhizopus by Oi et al. in Agr. Biol. Chem. 33, 729-38 (1969); and for Mucor by Somkuti et al. in Appl. Microbiol. 16, 617-19 (1968). All the lipases mentioned have a higher specificity for medium to long chain fatty acids, such as Aspergillus or Rhizopus, or for medium chain fatty acids, as is the case for Mucor, compared to short chain fatty acids, such as C4- to C6-fatty acids.

[0008] Microbial lipases are also available from Yarrowia, such as Yarrowia lipolytica. Several documents disclose the preference of lipases from Yarrowia for medium- to long-chain fatty acids. For example, Kamoun et al. 2015 described a higher specificity for trioctanoin compared to tributyrin at pH 7.5 for Lip8; Aloulou et al. 2007 disclosed a higher specificity for trioctanoin compared to tributyrin at pH 6 for Lip2; Sheng, J., et al. 2012 reported that LipY prefers C8-C 12 It is noted to have the highest activity towards fatty acids.

[0009] US2005059130 (or EP 2 290 059) also describes microbial lipases from Humicola lanuginosa, Fusarium oxysporum, or Rhizomucor miehei (Examples 10, 11, and 12, respectively), in which the substrate specificity of the wild-type enzyme is modified by altering the amino acid sequence of the lipase, thereby obtaining variants with enhanced specificity for short-chain fatty acids. EP3 081 644 also discloses a modified lipase from Candida cylindracea with higher specificity for short-chain fatty acids. However, none of the engineered lipases from these microorganisms are naturally occurring lipases with a higher specificity for short-chain fatty acids compared to medium- to long-chain fatty acids.

[0010] The above illustrates the difficulty of obtaining wild-type microbial lipases with higher specificity for short-chain fatty acids, especially C4-fatty acids, and lower specificity for medium and / or long-chain fatty acids, especially lower specificity for long-chain fatty acids, which, however, lead to reduced soapiness and increased butyric flavor in food products such as dairy or cheese, and thus can replace currently available microbial lipases. Summary of the Invention [Problem to be solved by the invention]

[0011] The present invention provides novel wild-type microbial lipolytic enzymes suitable for use in the food industry, preferably the dairy industry, more preferably the cheese industry. Surprisingly, the enzymes disclosed herein have a higher specificity for short-chain fatty acids, particularly C4- and / or C6-fatty acids, when compared to known microbial lipolytic enzymes under the same conditions, particularly for the same enzyme dosage, and in the same fat matrix, meaning that these enzymes release, produce or create more short-chain fatty acids than the known microbial enzyme(s). This is completely unexpected, as until now wild-type microbial lipolytic enzymes have been specific for medium or long-chain fatty acids and are not directed towards short-chain fatty acids, preferably C4- and / or C6-fatty acids. [Means for solving the problem]

[0012] The present invention: has at least 90% identity to 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 or SEQ ID NO:11, preferably SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, more preferably SEQ ID NO:1, SEQ ID NO:3 or SEQ ID NO:11, even more preferably SEQ ID NO:1, or having at least 95%, 96%, 97%, 98%, 99% identity to 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 or SEQ ID NO:11, preferably SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, more preferably SEQ ID NO:1, SEQ ID NO:3 or SEQ ID NO:11, even more preferably SEQ ID NO:1, containing an amino acid sequence, or wherein the sequence is 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 or SEQ ID NO: 11, preferably SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 9 or SEQ ID NO: 11, more preferably SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 11, even more preferably SEQ ID NO: 1, Concerning lipase.

[0013] The present invention also provides: has at least 90% identity to 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 or SEQ ID NO:11, preferably SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, more preferably SEQ ID NO:1, SEQ ID NO:3 or SEQ ID NO:11, even more preferably SEQ ID NO:1, or having at least 95%, 96%, 97%, 98%, 99% identity to 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 or SEQ ID NO:11, preferably SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, more preferably SEQ ID NO:1, SEQ ID NO:3 or SEQ ID NO:11, even more preferably SEQ ID NO:1, containing an amino acid sequence, or wherein the sequence is 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 or SEQ ID NO:11, preferably SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, more preferably SEQ ID NO:1, SEQ ID NO:3 or SEQ ID NO:11, even more preferably SEQ ID NO:1, and Here, the lipase is preferably at a pH of less than 7, preferably at a pH of 6.6 to 6.8, preferably at a pH of less than 6, preferably at a pH of 3.8 to 5.6, more preferably at a pH of 4.4 to 5.4, even more preferably at a pH of 4.6 to 5.2, and / or at a temperature of preferably less than 40°C, less than 30°C, or less than 20°C, preferably less than 15°C, more preferably less than 10°C, even more preferably at a temperature of 5 to 8°C. 10 - have a higher specificity for the release of C4-fatty acids from milk fat and / or milk compositions containing other fats when compared with the release of fatty acids; It also relates to lipase.

[0014] The present invention also provides: has at least 90% identity to 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 or SEQ ID NO:11, preferably SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, more preferably SEQ ID NO:1, SEQ ID NO:3 or SEQ ID NO:11, even more preferably SEQ ID NO:1, or having at least 95%, 96%, 97%, 98%, 99% identity to 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 or SEQ ID NO:11, preferably SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, more preferably SEQ ID NO:1, SEQ ID NO:3 or SEQ ID NO:11, even more preferably SEQ ID NO:1, containing an amino acid sequence, or wherein the sequence is 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 or SEQ ID NO: 11, preferably SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 9 or SEQ ID NO: 11, more preferably SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 11, even more preferably SEQ ID NO: 1, Concerning lipase.

[0015] The present invention also provides: has at least 90% identity to 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 or SEQ ID NO:11, preferably SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, more preferably SEQ ID NO:1, SEQ ID NO:3 or SEQ ID NO:11, even more preferably SEQ ID NO:1, or having at least 95%, 96%, 97%, 98%, 99% identity to 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 or SEQ ID NO:11, preferably SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, more preferably SEQ ID NO:1, SEQ ID NO:3 or SEQ ID NO:11, even more preferably SEQ ID NO:1, containing an amino acid sequence, or wherein the sequence is 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 or SEQ ID NO:11, preferably SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, more preferably SEQ ID NO:1, SEQ ID NO:3 or SEQ ID NO:11, even more preferably SEQ ID NO:1, and The lipase is preferably at a pH of 3.8 to 5.6, more preferably at a pH of 4.4 to 5.4, even more preferably at a pH of 4.6 to 5.2, and / or at a temperature of preferably less than 20°C, preferably less than 15°C, more preferably less than 10°C, even more preferably at 5 to 8°C. 10-When compared with respect to the release of fatty acids, when compared with the release of C8-fatty acids, when compared with the release of C6-fatty acids, 12 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 18:0 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 16 -When compared to the release of fatty acids, or C8-fatty acids ~ C 18:2 -When compared to the release of fatty acids, C 10 -Fatty acid~C 18:1 -When compared to the release of fatty acids, C 12 -Fatty acid~C 18:0 -relative to the release of fatty acids, or C 14 -Fatty acid~C 16:0 The present invention also relates to lipases that have a higher specificity for the release of C4-fatty acids from milk fat and / or dairy compositions containing other fats when compared to the release of C5-fatty acids.

[0016] In preferred embodiments, the lipases disclosed herein—SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11—are isolated microbial lipases, recombinant microbial lipases, or synthetic microbial lipases.

[0017] Although the present invention relates to wild-type, non-recombinant, microbial lipolytic enzymes, the skilled artisan will appreciate that one or more modifications do not interfere with the activity of the enzyme and preferably do not involve the C of its sequence. 4:0 It is recognized that modifications can be made to any of the amino acid sequences of the enzymes disclosed herein without altering their fatty acid preference. Such modifications can be additions, substitutions, or deletions. For example, altering the N- or C-terminus of any one of the sequences disclosed herein may not result in a different sequence, but may alter the fatty acid preference of the altered sequence relative to the unaltered sequence. These modifications are also contemplated herein.

[0018] The present invention also relates to isolated, recombinant or synthetic DNA sequences, wherein the isolated DNA sequence, recombinant DNA sequence or synthetic DNA is selected from sequences having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22 or SEQ ID NO:23. In a preferred embodiment, the DNA sequence may further comprise a signal peptide sequence.

[0019] The present invention further relates to vectors, such as cloning and / or expression vectors, comprising isolated, recombinant or synthetic DNA sequences encoding the lipases 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 or SEQ ID NO:11 disclosed herein, or having at least 90% sequence identity to 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 or SEQ ID NO:11, or having at least 95%, 96%, 97%, 98% or 99% sequence identity to 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 or SEQ ID NO:11. The DNA sequence may be SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, or SEQ ID NO:23, or a sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, or SEQ ID NO:23. The sequence is cloned into a vector using standard cloning techniques. The choice of cloning and / or expression vector depends on the host cell, as the cloning and / or expression vector must be compatible with the host cell.

[0020] The present invention also relates to a nucleic acid sequence comprising 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, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22 or SEQ ID NO:23 or having at least 90% sequence identity to 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 or SEQ ID NO:11 or a nucleic acid sequence .... The present invention also relates to a host cell, preferably a genetically modified host cell, comprising a sequence having at least 95%, 96%, 97%, 98% or 99% sequence identity to 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 or SEQ ID NO:11, or comprising a sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity to SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22 or SEQ ID NO:23.

[0021] The invention also relates to methods for propagating, transforming or transfecting said vectors in suitable host cells and under conditions conducive to expression of the polypeptide or overexpression of the polypeptide. Suitable host cells for use in the present invention may be any host cell in which recombinant DNA can be replicated and enzyme-encoding genes can be expressed. Preferably, the host cell may be a mammalian cell, an insect cell, a microbial cell, such as a bacterial cell, or a fungal cell, including a yeast cell.

[0022] Examples of suitable yeasts may be Saccharomyces cerevisiae; Saccharomyces spp., such as Candida cylindracea or Pichia pastoris, Schizosaccharomyces spp., Candida spp., Pichia spp., Kluyveromyces spp.

[0023] Examples of suitable fungal cells include filamentous fungal cells, which may be selected from Fusarium spp., such as Fusarium oxysporium, Aspergillus oryzae, or Aspergillus niger, Aspergillus spp., and Trichoderma spp.

[0024] Examples of suitable bacteria are gram-negative bacteria such as Escherichia coli, or other bacteria such as Bacillus subtilis, Bacillus licheniformis, Bacillus lentus, Bacillus brevis, Bacillus stearothermophilus, Bacillus alkalophilus, Bacillus amyloliquefaciens, Bacillus coagulans, Bacillus circulans, Bacillus lautus, Bacillus megaterium, Bacillus The Gram-positive bacterium is selected from the group of Bacillus species, such as Bacillus megaterium, Bacillus thuringiensis, Streptomyces lividans or Streptomyces murinus, Streptomyces species, Corynebacterium species. Transformation of bacteria may be carried out, for example, by using competent cells in a manner known in the art, or by other methods known in the art.

[0025] In a preferred embodiment, the host cell is selected from the group consisting of Pichia pastoris, Saccharomyces species, Saccharomyces cerevisiae, Schizosaccharomyces species, Candida species, Candida cylindracea, Kluyveromyces species, Fusarium species, Fusarium oxysporium, Aspergillus subsp. Aspergillus oryzae or Aspergillus niger, Trichoderma species, Escherichia coli or Bacillus species, Bacillus subtilis, Bacillus licheniformis, Bacillus lentus, Bacillus brevis, Bacillus stearothermoides ... philus, Bacillus alkalophilus, Bacillus amyloliquefaciens, Bacillus coagulans, Bacillus circulans, Bacillus lautus, Bacillus megaterium, Bacillus thuringiensis, the group of Streptomyces species, Streptomyces lividans or Streptomyces murinus, the group of Corynebacterium species, preferably Pichia pastoris, Aspergillus oryzae or Aspergillus niger, or Bacillus subtilis, more preferably Pichia pastoris or Bacillus subtilis.

[0026] The present invention also relates to a lipase consisting of an amino acid sequence having at least 90%, at least 95%, at least 96%, or at least 97%, at least 98%, at least 99% or 100% identity to 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 or SEQ ID NO:11, preferably having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11; wherein the lipase is expressed in Pichia pastoris and is isolated from C. 10 - compared to the release of C8-fatty acids, compared to the release of C6-fatty acids, 12 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C18:0 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 16 -When compared to the release of fatty acids, or C8-fatty acids ~ C 18:2 -When compared to the release of fatty acids, C 10 -Fatty acid~C 18:1 -When compared to the release of fatty acids, C 12 -Fatty acid~C 18:0 -When compared to the release of fatty acids, C 14 -Fatty acid~C 16:0 -When compared to the release of fatty acids, C 16 ~C 18 The present invention has a higher specificity for the release of C4-fatty acids from milk fat and / or dairy compositions containing other fats when compared to the release of C5-fatty acids from milk fat and / or dairy compositions containing other fats.

[0027] The present invention also relates to a method for preparing a food product, comprising: - using or adding a lipase comprising an amino acid sequence which has at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to 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 or SEQ ID NO:11, preferably having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, preferably wherein the food product has a higher content, amount or concentration of short chain fatty acids, such as C4- and / or C6-fatty acids compared to the content, amount or concentration of medium to long chain fatty acids; or

[0028] - using or adding a lipase encoded by a DNA sequence selected from a sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22 or SEQ ID NO:23, preferably having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:20, SEQ ID NO:22 or SEQ ID NO:23, and preferably wherein the food product has a higher content, amount or concentration of short chain fatty acids, such as C4- and / or C6-fatty acids compared to the content, amount or concentration of medium to long chain fatty acids.

[0029] In a preferred embodiment, the food product may be a cheese product, processed cheese, cheese-like product, or enzyme-treated cheese, or a dairy product such as butter, yogurt, cream, seasonings, etc.

[0030] In a preferred embodiment, the dairy product is a cheese product, preferably wherein the cheese product is Pecorino, Provolone, Parmesan, Grana Padano, Parmigiano-Reggiano, Romano, Chester, Dumbo, Manchego, Saint Paulin, Cheddar, Monterey, Colby, Edam, Gouda, Muenster, Swiss-type, Gruyère, Emmental; curd cheese such as feta; pasta filata cheese such as mozzarella or queso fresco; fresh cheese such as ricotta, cream cheese, Neuchâtel or cottage cheese; cream cheese, white-veined cheese such as brie or camembert, blue-veined cheese such as gorgonzola or Danish blue cheese; and processed cheese, preferably feta or provolone.

[0031] In another embodiment, the food product may be a dairy alternative product, preferably a dairy alternative cheese or a vegan cheese, more preferably wherein the lipase is 10 - compared to the release of C8-fatty acids, compared to the release of C6-fatty acids, 12 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 18:0 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 16 -When compared to the release of fatty acids, or C8-fatty acids ~ C 18:2 -When compared to the release of fatty acids, C 10 -Fatty acid~C 18:1 -When compared to the release of fatty acids, C 12 -Fatty acid~C 18:0 -When compared to the release of fatty acids, C 14 -Fatty acid~C 16:0 -relative to the release of fatty acids, or C 16 ~C 18 It has a higher specificity for the release of C4-fatty acids from lipid-containing compositions such as vegetable fats or from water-in-oil emulsions when compared to the release of C5-fatty acids from water-in-oil emulsions or from lipid-containing compositions such as vegetable fats.

[0032] In a preferred embodiment, cheese flavor or cheese strength is enhanced and / or improved by applying a lipase of the present invention. The enhancement or improvement of cheese flavor or cheese strength can be applied to cheese itself, or to processed cheese, cheese-like products, or enzyme-treated cheese, or even to dairy alternative cheeses such as vegan cheese.

[0033] In a preferred embodiment, the lipases disclosed herein may be added to an intermediate product, for example, in the process of making a food product, or to a raw material such as milk, particularly milk obtained from cows, sheep, or goats, among others.

[0034] The present invention also relates to the use of a lipase comprising an amino acid sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to 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 or SEQ ID NO:11, preferably having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, preferably wherein the food product has a higher content, amount or concentration of short-chain fatty acids, such as C4- and / or C6-fatty acids, as compared to the content, amount or concentration of medium- to long-chain fatty acids, and wherein the food product is present as a dairy product, a cheese product, a processed cheese, a cheese-like product, an enzyme-treated cheese, or as butter, yogurt, cream, seasoning.

[0035] In a preferred embodiment, the lipase is used to produce food products, in particular dairy products or cheeses such as Pecorino, Provolone, Parmesan, Grana Padano, Parmigiano-Reggiano, Romano, Chester, Dumbo, Manchego, Saint Paulin, Cheddar, Monterey, Colby, Edam, Gouda, Muenster, Swiss-type, Gruyère, Emmental; curd cheeses such as Feta; pasta filata cheeses such as Mozzarella and Queso Fresco; fresh cheeses such as Ricotta, cream cheese, Neuchâtel or cottage cheese; cream cheese, white-veined cheeses such as Brie or Camembert, blue-veined cheeses such as Gorgonzola or Danish blue cheese, preferably Feta or Provolone; or processed cheese.

[0036] The present invention also provides a method for producing fatty acids containing more C6-, C8-, C 10 -Fatty acids, C 12 -Fatty acids, C 14 -Fatty acids, C16 -Fatty acids, C 18:0 -Fatty acids, C 18:1 -Fatty acids, C 18:2 -fatty acids, or C 18:3 The present invention also relates to the use of a lipase to produce or release more C4-fatty acids from a dairy composition comprising milk fat and / or other lipids compared to producing or releasing C4-fatty acids, wherein the lipase comprises an amino acid sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to 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 or SEQ ID NO:11, preferably having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11.

[0037] The present invention also provides a method for the preparation of C8-fatty acids, C 10 -Fatty acids, C 12 -Fatty acids, C 14 -Fatty acids, C 16 -Fatty acids, C 18:0 -Fatty acids, C 18:1 -Fatty acids, C 18:2 -fatty acids, or C 18:3 The present invention also relates to the use of a lipase for producing or releasing more C4- and / or C6-fatty acids in a food product compared to producing or releasing -fatty acids, wherein the lipase comprises an amino acid sequence that has at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to 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 or SEQ ID NO:11, preferably having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11,

[0038] Preferably wherein the food product is cheese, or a processed cheese, enzyme treated cheese, or cheese-like product, or a dairy product selected from butter, yogurt, cream, seasonings, or Preferably herein, the food product is a dairy alternative such as a dairy alternative cheese or a vegan cheese.

[0039] In one embodiment, the food product is a dairy product selected from cheese, and wherein the cheese is feta, provolone, pecorino, parmesan, grana padano, parmigiano reggiano, romano, chester, dambo, manchego, saint paulin, cheddar, monterey, colby, edam, gouda, muenster, swiss-type cheese, gruyere, or emmental; or pasta filata cheese such as mozzarella or queso fresco; or fresh cheese such as ricotta, cream cheese, neuchâtel, or cottage cheese; or cream cheese; or white mould cheese such as brie or camembert; or blue mould cheese such as gorgonzola or danish blue, preferably feta or provolone.

[0040] Furthermore, the present invention relates to a food product obtainable by using a lipase 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 or SEQ ID NO:11, preferably wherein the food product is a cheese product, processed cheese, cheese-like product or a dairy product such as butter, yogurt, cream, seasoning, more preferably the dairy product is Pecorino, Provolone, Parmesan, Grana Padano, Parmigiano-Reggiano, Romano, Chester, Dumbo, Manchego , Saint Paulin, Cheddar, Monterey, Colby, Edam, Gouda, Muenster, Swiss-type, Gruyère, Emmental; curd cheeses such as feta; pasta filata cheeses such as mozzarella and queso fresco; fresh cheeses such as ricotta, cream cheese, Neuchâtel or cottage cheese; cream cheese, white mold cheeses such as brie and camembert, blue mold cheeses such as gorgonzola and Danish blue cheese; and processed cheeses, preferably feta or provolone.

[0041] The present invention also relates to a food product comprising a lipase, wherein the lipase comprises an amino acid sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to 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 or SEQ ID NO:11, preferably having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11; and wherein the food product is C 10 -fatty acids, or C6-fatty acids, C8-fatty acids, C 12 -Fatty acids, C 14 -Fatty acids, C 16 -Fatty acids, C 18:1 -Fatty acids, C 18:2-fatty acid or C 18:3 -contains more C4-fatty acids than other fatty acids selected from the fatty acids; and / or wherein the food product is C 10 -fatty acids, or C8-fatty acids, C 12 -Fatty acids, C 14 -Fatty acids, C 16 -Fatty acids, C 18:1 -Fatty acids, C 18:2 -fatty acid or C 18:3 -contains more C6-fatty acids than other fatty acids selected from fatty acids.

[0042] In some embodiments, the food product is cheese, processed cheese, cheese-like products, enzyme-treated cheese, or a dairy product selected from butter, yogurt, cream, and seasonings, preferably wherein the cheese is feta cheese, provolone cheese, pecorino cheese, parmesan cheese, grana padano cheese, parmigiano-reggiano cheese, romano cheese, chester cheese, dambo cheese, manchego cheese, saint paulin cheese, cheddar cheese, monterey cheese, colby cheese, edam cheese, gouda cheese, muenster cheese, swiss-type cheese, gruyere cheese, or emmental cheese; or pasta filata cheese such as mozzarella or queso fresco cheese; or fresh cheese such as ricotta, cream cheese, neuchâtel, or cottage cheese; or cream cheese; or white-veined cheese such as brie or camembert cheese; or blue-veined cheese such as gorgonzola or danish blue cheese; more preferably feta cheese or provolone cheese. In another embodiment, the food product is a dairy alternative, such as a dairy alternative cheese or a vegan cheese.

[0043] The present invention also relates to a lipase according to 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 or SEQ ID NO:11 and / or a DNA sequence according to SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22 or SEQ ID NO:23 and / or a DNA sequence according to 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, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22 or SEQ ID NO:23. 17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22 or SEQ ID NO:23 and / or a host cell comprising any of the sequences 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, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22 or SEQ ID NO:23, preferably wherein the composition is for preparing a food product such as a dairy product or cheese.

[0044] definition In the context of the present invention, the terms "short-chain fatty acids", "medium-chain fatty acids", "long-chain fatty acids", "FFA", "lipase", "lipolytic enzyme", "microbial lipase", "microbial lipolytic enzyme", "lipase with higher specificity for the release of short-chain fatty acids", "lipase with higher specificity for the release of C4-fatty acids", "lipase with higher specificity for the release of C6-fatty acids", "lipase with higher specificity for the release of medium and / or long-chain fatty acids", "lipase activity", "lipase dosage", "wild-type", "synthetic sequence", "isolated sequence", "recombinant sequence", "mature protein sequence", "expression vector", "sequence identity", "food product", "dairy product", "cheese", "cheese product", "processed cheese", "enzyme-treated cheese", "cheese-like product", "improvement factor (IF)", "control microbial lipase", "signal peptide", "propeptide" have the meanings explained below.

[0045] The term "short chain fatty acid" means a fatty acid containing 4 to 6 carbons. Examples of short chain fatty acids are: butyric acid (butanoic acid; C4), valeric acid (pentanoic acid; C5) or caproic acid (hexanoic acid; C6).

[0046] The term "medium chain fatty acids" refers to fatty acids containing 8 to 12 carbons. Examples of medium chain fatty acids are: caprylic acid (octanoic acid; C8), pelargonic acid (nonanoic acid; C9), capric acid (decanoic acid; C10), 10 ), undecylic acid (undecanoic acid; C 11 ) or lauric acid (dodecanoic acid; C 12 )

[0047] The term "long-chain fatty acid" refers to a fatty acid containing 14 to 18 or more carbon atoms. Examples of long-chain fatty acids include: myristic acid (tetradecanoic acid; C 14 ), pentadecanoic acid (C 15 ), palmitic acid (hexadecanoic acid; C 16 ), margaric acid (heptadecanoic acid; C 17 ), stearic acid (octadecanoic acid; C 18 )

[0048] The term "FFA" stands for free fatty acids. The terms "lipase" and "lipolytic enzyme" are used interchangeably.

[0049] The terms "microbial lipase" and "microbial lipolytic enzyme" are used interchangeably and, as used herein, refer to lipases or lipolytic enzymes expressed by naturally occurring microorganisms found in nature. Furthermore, the microorganism may be a filamentous fungus and / or a non-filamentous fungus. Thus, the lipase, microbial lipase, or microbial lipolytic enzyme may be a filamentous fungal lipase or a non-filamentous fungal lipase. The term "lipase with higher specificity for the release of short-chain fatty acids" from a dairy composition containing milk fat and / or other lipids means that the lipase preferably hydrolyzes the ester bonds of lipids in triglycerides to produce more C4-fatty acids and / or more C6-fatty acids, rather than medium- and / or long-chain fatty acids.

[0050] The term "lipase with higher specificity for the release of C4-fatty acids" from dairy compositions containing milk fat and / or other lipids means that said lipase preferably hydrolyzes the ester bonds of lipids in triglycerides to give more C4-fatty acids rather than other types of fatty acids such as medium and / or long chain fatty acids. 10 The term "lipase with higher specificity for the release of C4-fatty acids" from dairy compositions containing milk fat and / or other lipids means that said lipase preferably hydrolyzes the ester bonds of lipids of triglycerides to release C4-fatty acids compared to the release of C4-fatty acids. 10 This means that more C4-fatty acids are produced rather than C8-fatty acids, C6-fatty acids, C 12 -fatty acid or C 18:2 -Fatty acids, C 14 -Fatty acids, C 18:0 -Fatty acids, C 18:1 -fatty acid or C 16The term "lipase with higher specificity for the release of C4-fatty acids" from dairy compositions containing milk fat and / or other lipids means that said lipase preferably hydrolyzes the ester bonds of lipids in triglycerides to release C8-fatty acids, C6-fatty acids, C8-fatty acids, C9-fatty acids, C10-fatty acids, C11-fatty acids, C12-fatty acids, C13-fatty acids, C14-fatty acids, C15-fatty acids, C16-fatty acids, C17-fatty acids, C18-fatty acids, C19-fatty acids, C20-fatty acids, C210-fatty acids, C22-fatty acids, C23-fatty acids, C24-fatty acids, C25-fatty acids, C26-fatty acids, C27-fatty acids, C28-fatty acids, C29-fatty acids, C30-fatty acids, C3 12 -Fatty acids, C 18:2 -Fatty acids, C 14 -Fatty acids, C 18:0 -Fatty acids, C 18:1 -fatty acid or C 16 This means that the molar fraction of C4-fatty acids is higher than that of medium and / or long chain fatty acids under the same test conditions, and preferably the molar fraction of C4-fatty acids is higher than that of C6-fatty acids, C8-fatty acids, C 10 -Fatty acids, C 12 -Fatty acids, C 14 -Fatty acids, C 16 -Fatty acids, C 18:0 -Fatty acids, C 18:1 -fatty acid or C 18:2 -higher compared to the mole fraction of other fatty acids such as fatty acids.

[0051] The term "lipase with higher specificity for the release of C4-fatty acids" from a dairy composition containing milk fat and / or other lipids also means that the lipase preferably hydrolyzes the ester bonds of lipids in triglycerides to produce more C4-fatty acids than the control microbial lipase. In particular, the mole fraction of C4-fatty acids for the test lipase is higher than the mole fraction of C4-fatty acids for the control microbial lipase.

[0052] The term "lipase with higher specificity for the release of C6-fatty acids" from a dairy composition containing milk fat and / or other lipids means that the lipase preferably hydrolyzes the ester bonds of lipids in triglycerides to produce more C6-fatty acids rather than other types of fatty acids such as medium and / or long chain fatty acids. 10The term "lipase with higher specificity for the release of C6-fatty acids" from dairy compositions containing milk fat and / or other lipids means that said lipase preferably hydrolyzes the ester bonds of lipids in triglycerides to release C6-fatty acids compared to the release of C6-fatty acids. 10 This means that more C6-fatty acids are produced rather than C8-fatty acids. 12 -fatty acid or C 18:2 -Fatty acids, C 14 -Fatty acids, C 18:0 -Fatty acids, C 18:1 -fatty acid or C 16 The term "lipase with higher specificity for the release of C6-fatty acids" from dairy compositions containing milk fat and / or other lipids means that said lipase preferably hydrolyzes the ester bonds of lipids of triglycerides to release C8-fatty acids, C9-fatty acids, C10-fatty acids, C12-fatty acids, C13-fatty acids, C14-fatty acids, C15-fatty acids, C16-fatty acids, C17-fatty acids, C18-fatty acids, C19-fatty acids, C20-fatty acids, C211-fatty acids, C221-fatty acids, C23-fatty acids, C24-fatty acids, C25-fatty acids, C26-fatty acids, C27-fatty acids, C28-fatty acids, C29-fatty acids, C30-fatty acids, C31-fatty acids, C32-fatty acids, C33-fatty acids 12 -Fatty acids, C 18:2 -Fatty acids, C 14 -Fatty acids, C 18:0 -Fatty acids, C 18:1 -fatty acid or C 16 This means that the molar fraction of C6-fatty acids is higher than that of medium and / or long chain fatty acids under the same test conditions, and preferably the molar fraction of C6-fatty acids is higher than that of C8-fatty acids, C 10 -Fatty acids, C 12 -Fatty acids, C 14 -Fatty acids, C 16 -Fatty acids, C 18:0 -Fatty acids, C 18:1 -fatty acid or C 18:2 -higher compared to the mole fraction of other fatty acids such as fatty acids.

[0053] The term "lipase with higher specificity for the release of C6-fatty acids" from a dairy composition containing milk fat and / or other lipids also means that the lipase preferably hydrolyzes the ester bonds of lipids in triglycerides to produce more C6-fatty acids than the control microbial lipase. In particular, the mole fraction of C6-fatty acids for the test lipase is higher than the mole fraction of C4-fatty acids for the control microbial lipase.

[0054] The term "lipase with higher specificity for the release of medium and / or long chain fatty acids" from dairy compositions containing milk fat and / or other lipids means that the lipase preferably hydrolyzes the ester bonds of lipids in triglycerides to produce C8-C fatty acids rather than C4- and / or C6-fatty acids. 18 -fatty acids. In particular, the C8 to C6 18 - The mole fraction of fatty acids is C8 to C6 for the control microbial lipase. 18 -higher compared to the mole fraction of fatty acids.

[0055] In the context of the present invention, the following terms are used: "C4-fatty acid" and "C 4:0 -fatty acid”; “C6-fatty acid” and “C 6:0 -fatty acids"; "C8-fatty acids" and "C 8:0 -fatty acid”; “C 10 -fatty acids" and "C 10:0 -fatty acid”; “C 12 -fatty acids" and "C 12:0 -fatty acid”; “C 14 -fatty acids" and "C 14:0 -fatty acids" are interchangeable. 16 The term "C-fatty acid" 16:0 -Fatty acids and C 16:1 - May contain fatty acids. 18 The term "C-fatty acid" 18:0 -Fat, C 18:1 -Fatty acids and C 18:3 -May contain fat.

[0056] The terms "lipase activity" and "lipase dosage" are used interchangeably. Lipase activity, for example, can be measured by the International Dairy Federation (IDF) method for both commercial lipases and the enzymes disclosed herein (SEQ ID NOS: 1-11) in relation to their dosage in cheese and cream. The IDF method primarily corresponds to the international standard ISO 13082:2011, first edition, November 15, 2011, IDF 218:2011 (also designated ISO 13082|218). However, in the present invention, the term "LFU / L of milk" is used. The term "LFU / L of milk" refers to the concentration or amount of enzyme or enzyme dosage per liter of milk used in producing dairy products such as cheese. In the context of the present invention, "LFU / L of milk" refers to the number of pre-gastric units of lipase per liter of milk. One LFU is defined as the amount of lipase activity that releases butyric acid at a rate of 1.25 μmol / min under the test conditions. Furthermore, lipase activity may also be measured as explained in the examples below. Finally, there are other methods for measuring lipase activity disclosed in prior art documents (such as EP 3 081 644, EP 2 254 996 or EP 1 776 455). What is relevant, however, is that within the same example or comparative example, lipase activity is measured in the same way for all lipases under analysis, including the reference lipase.

[0057] In the context of the present invention, the term "wild-type" lipase means that the sequence is not mutated compared to the mature protein sequence endogenously produced (after co-translational and / or post-translational cleavage events), i.e., the lipase does not contain amino acid deletions, additions, or substitutions. The wild-type lipase of the present invention may be encoded by a codon-optimized polynucleotide for heterologous expression and may also contain a non-endogenous signal and / or propeptide selected for expression in a given host. Furthermore, in the present invention, the lipase may also be synthetically obtained or recombinantly engineered using methods well known in the art to regenerate the wild-type lipase.

[0058] The term "synthetic sequence" or variations thereof refers to a sequence prepared by in vitro chemical or enzymatic synthesis. It includes, but is not limited to, sequences made using optimal codon usage for the host organism. The term applies to synthetic microbial lipases or synthetic DNA sequences.

[0059] The term "isolated sequence" or variations thereof refers to a sequence that is at least substantially free from at least one other component with which the sequence is actually associated and found in nature. Alternatively, "isolated sequence" can refer to a sequence that has been removed from its natural environment. For example, a sequence that has been produced recombinantly in a host organism would be considered isolated for the purposes of the present invention, and purification methods are known in the art, as are naturally occurring or recombinant sequences that have been substantially purified by any suitable technique per se, e.g., purification methods. The term applies to an isolated microbial lipase or an isolated DNA sequence.

[0060] The term "recombinant sequence" or variations thereof means a sequence prepared using recombinant DNA techniques and is within the ability of one skilled in the art and is described in the literature, e.g., J. Sambrook, E.F. Fritsch, and T. Maniatis, 1989, Molecular Cloning: A Laboratory manual, 2nd edition, books 1-3, Cold Spring Harbor Laboratory Press. The term applies to recombinant microbial lipases or recombinant DNA sequences.

[0061] As used herein, the term "mature protein sequence" refers to a sequence that has lipolytic activity in its final form after translation and co- and / or post-translational cleavage events or modifications such as N-terminal processing, C-terminal cleavage, glycosylation, phosphorylation, among other events or modifications.

[0062] The term "expression vector" means a construct capable of in vivo or in vitro expression. Expression vectors can be integrated, in particular stably integrated, into the genome of a suitable host organism.

[0063] The term "sequence identity" describes a quantitative measure of similarity between two amino acid sequences or two nucleotide sequences. For purposes of the present invention, the degree of sequence identity between two amino acid sequences is based on aligning both sequences with the blastp suite provided by the National Center for Biotechnology Information (NCBI) at https: / / blast.ncbi.nlm.nih.gov, which applies standard parameter settings (matrix: BLOSUM62, gap cost: yes: 11, extension: 1, condition composition score matrix adjustment), followed by quantification of identical amino acid pairs at identical positions throughout the aligned amino acid sequences.

[0064] The term "food product" refers to any dairy product intended to be used as food, including, but not limited to, cheese, milk, skim milk, sour milk, buttermilk, condensed milk, spreads, margarine, yogurt, ice cream, milk powder, butter, and dulce de leche.

[0065] The term "dairy product" is intended to include any food product made using milk or a dairy product, including, but not limited to, cheese, milk, skim milk, sour milk, buttermilk, condensed milk, spreads, margarine, yogurt, ice cream, milk powder, butter, and dulce de leche.

[0066] The terms "cheese" and "cheese products" are used interchangeably. The term "cheese" is understood to encompass any cheese, including, but not limited to, hard cheeses such as Pecorino, Provolone, Parmesan, Grana Padano, Parmigiano-Reggiano, Romano, Chester, Dumbo, Manchego, Saint Paulin, Cheddar, Monterey, Colby, Edam, Gouda, Muenster, Swiss-type, Gruyère, and Emmental; curd cheeses such as feta; pasta filata cheeses such as mozzarella and queso fresco; fresh cheeses such as ricotta, cream cheese, Neuchâtel, or cottage cheese; cream cheese, or white-mold cheeses such as Brie or Camembert, or blue-mold cheeses such as Gorgonzola or Danish blue cheese; and processed cheese, enzyme-modified cheese (EMC), or cheese-like products.

[0067] The term "processed cheese" refers to cheese or cheese analogs preferably produced by heating and emulsifying the cheese, such as with emulsifying salts (e.g., phosphates and citrates). The process may further include the addition of spices / condiments.

[0068] The term "enzyme-treated cheese" or "EMC" is understood to mean cheese curd that has been treated with an enzyme to produce a rich cheese flavor component that can have about 15 to 30 times the flavor intensity of natural cheese. EMC is available as a paste or dried to form a powder and is used to impart cheese flavor notes to products such as processed cheese or cheese analogs, cheese powders, soups, sauces, dips, crackers, salad dressings, and snack food coatings.

[0069] The term "cheese-like product" is understood as a cheese-like product that comprises, as part of its composition, a lipid, such as, for example, a dairy fat (e.g., cream or butter) or a vegetable oil, and that further comprises, as part of its composition, one or more non-dairy ingredients, such as, for example, a vegetable ingredient (e.g., a vegetable protein or vegetable oil). In the context of the present invention, "cheese-like product" includes dairy-alternative cheeses, also known as vegan cheeses, in which the lipid used to make the dairy-alternative cheeses is, instead of dairy fat, a vegetable oil or an emulsion, such as a water-in-oil emulsion.

[0070] The term "improvement factor (IF)" is defined as the ratio obtained when the enzymatic specificity of the test lipase for the release of short-chain fatty acids is divided by the enzymatic specificity of the control lipase, preferably the control microbial lipase, for the release of short-chain fatty acids, or the ratio obtained when the enzymatic specificity of the test lipase for the release of medium-chain fatty acids is divided by the enzymatic specificity of the control lipase, preferably the control microbial lipase, for the release of medium-chain fatty acids. If the ratio is 1, then the test lipase has the same preference or specificity for short-chain fatty acids as the control sequence. If the ratio is lower than 1, then the test lipase has a lower preference or specificity for short-chain fatty acids compared to the control lipase, preferably the control microbial lipase. If the ratio is higher than 1, then the test lipase has a higher preference or specificity for short-chain fatty acids compared to the control. The enzymatic specificity for short-chain fatty acids is calculated by dividing the enzymatic activity for short-chain fatty acids C4 or C6 by the enzymatic activity for long-chain fatty acids C6. 16 ~C 18The improvement factor (IF) can also be determined by the ratio obtained by dividing the enzymatic specificity of the test lipase for the release of long-chain fatty acids by the enzymatic specificity of the control lipase, preferably the control microbial lipase, for the release of long-chain fatty acids. If the ratio is 1, then the test lipase has the same preference or specificity for long-chain fatty acids as the control lipase, preferably the control microbial lipase. If the ratio is lower than 1, then the test lipase has a lower preference or specificity for long-chain fatty acids compared to the control lipase. If the ratio is higher than 1, then the test lipase has a higher preference or specificity for long-chain fatty acids compared to the control. The enzymatic specificity for long-chain fatty acids is determined by the ratio of the long-chain fatty acid C 16 ~C 18 The enzyme activity for C4 or C6 short-chain fatty acids is obtained by dividing the enzyme activity for C4 or C6 short-chain fatty acids by the enzyme activity for C6 or C6 short-chain fatty acids.

[0071] The terms "control lipase," "control microbial lipase," "control lipolytic enzyme," or "control microbial lipolytic enzyme" refer to a commercially available microbial lipase derived from Rhizomucor miehei (Palatase®, Palatase® 20000 L available from Novozymes A / S), and are referred to herein as commercial microbial lipase or commercial microbial lipase A. Alternatively, other microbial lipases derived from Rhizomucor miehei can be used, such as SpiceIT® MR available from Chr. Hansen.

[0072] The term "signal peptide" relates to an amino acid sequence fused to the N-terminus of the polypeptide chain of an enzyme that is recognized by the secretory pathway of the enzyme-producing microorganism, which translocates the synthesized protein into the cell membrane or secretes it from the cell into the fermentation medium.

[0073] The term "propeptide" refers to a polypeptide chain fused to the N-terminus of the main peptide chain of an enzyme, which may support folding of the synthesized protein and / or contribute to its stability. The propeptide is cleaved during the maturation or activation of the enzyme. The propeptide acts as an N-terminal extension of the protein, distinct from the signal sequence required for the transport of the protein in or across a membrane or for its secretion into the extracellular medium. The presence of a propeptide sequence may lead to stabilization of the expression of the lipase and / or prevent its in vivo inactivation.

[0074] Amino acids are referred to herein by either their single letter code or their three letter code, both of which are well known to those skilled in the art.

[0075] In the context of the present invention, SEQ ID NOs: 1 to 11 represent 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, and SEQ ID NO: 11. Furthermore, in the context of the present invention, SEQ ID NOs: 12 to 23 represent SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, and SEQ ID NO: 23. DETAILED DESCRIPTION OF THE INVENTION

[0076] Detailed Description The present invention relates to non-animal, wild-type microbial lipases that meet vegetarian and / or kosher requirements while simultaneously exhibiting specificity and preference for cleavage of short chain fatty acids, preferably exhibiting higher specificity for C4- and / or C6-fatty acids rather than other types of fatty acids, or exhibiting higher specificity for C4- and / or C6-fatty acids compared to commercially available microbial lipases.

[0077] The objective of the present invention is to provide a microbial wild-type lipase that has a higher specificity for the release of C4 and / or C6 fatty acids from dairy compositions containing milk fat and / or other lipids, compared to the release of medium- and / or long-chain fatty acids, preferably compared to the release of long-chain fatty acids, without introducing any mutations (additions, deletions, and / or substitutions) into the lipase sequence, or a higher specificity for the release of C4 and / or C6 fatty acids from dairy compositions containing milk fat and / or other lipids, compared to the release of C4 and / or C6 fatty acids of commercially available microbial lipases. Thus, the present invention relates to improved (or alternative) microbial wild-type lipases that have specificity for short-chain fatty acids relative to conventional microbial lipases. Therefore, the lipases disclosed herein may be derived from microorganisms that naturally produce lipases. In this case, the lipase is a wild-type lipase.

[0078] The microbial lipases disclosed herein (SEQ ID NOs: 1-11) may be used to replace microbial lipases currently available on the market. [Example]

[0079] Example 1 - Cloning and Expression A nucleotide sequence 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, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11 is integrated into a target locus of an expression host, such as Pichia pastoris. A codon-optimized gene 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, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11 may or may not be used. The nucleotide sequence encoding SEQ ID NO:1 is, for example, SEQ ID NO:12 or SEQ ID NO:23. The nucleotide sequence encoding 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, or SEQ ID NO:11 may be, for example, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, or SEQ ID NO:22, respectively.

[0080] In one embodiment, Pichia pastoris is selected from the group consisting of Saccharomyces species, Saccharomyces cerevisiae, Schizosaccharomyces species, Candida species, Candida cylindracea, Kluyveromyces species, Fusarium species, Fusarium oxysporium, Aspergillus species, Aspergillus oryzae or Aspergillus niger, Trichoderma species, Escherichia coli or Bacillus species, Bacillus subtilis, Bacillus licheniformis, Bacillus lentus, Bacillus brevis, Bacillus stearothermophilus, Bacillus alkalophilus, Bacillus amyloliquefaciens, Bacillus coagulans, Bacillus circulans, Bacillus lautus, Bacillus megaterium, Bacillus thuringiensis, the group of Streptomyces species, Streptomyces lividans or Streptomyces murinus, the group of Corynebacterium species, preferably Aspergillus oryzae, Aspergillus niger, or Bacillus subtilis.

[0081] Nucleotide sequences 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, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11 may also be cloned into conventional cloning and expression vectors. Codon-optimized genes 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, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11 may or may not be used. The nucleotide sequence encoding SEQ ID NO:1 may be, for example, SEQ ID NO:12 or SEQ ID NO:23. The nucleotide sequence encoding 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, or SEQ ID NO:11 may be, for example, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, or SEQ ID NO:22, respectively.

[0082] The recombinant cloning and expression vectors may further be introduced into host cells, such as recombinant host cells, using standard transformation or transfection protocols known to those skilled in the art or described in the literature (e.g., J. Sambrook, E.F. Fritsch, and T. Maniatis, 1989, Molecular Cloning: A Laboratory manual, 2nd edition, books 1-3, Cold Spring Harbor Laboratory Press).

[0083] The microbial lipases disclosed herein (SEQ ID NOS: 1-11) were independently expressed as follows. Pichia pastoris was used as a host for lipase expression. Pichia pastoris was cultured on YPD agar plates (Invitrogen) under selection conditions (100 μg / mL Zeocin) at 30°C for 16 hours. A single colony from each YPD agar plate was used to inoculate 3 mL of YPD liquid medium (Invitrogen) containing 100 μg / mL Zeocin, followed by incubation at 30°C with 250 rpm shaking for 16 hours to perform seed fermentation. An OD of 0.2 was obtained. 600 Main fermentations were performed in a total volume of 4 mL by diluting the seed fermentation with BMGY medium (Invitrogen) containing 100 μg / mL Zeocin to 100 μg / mL, followed by incubation in 24-well culture plates at 30°C and 225 rpm for 3 days. The main fermentation samples were centrifuged to pellet the expression host cells, and the supernatant was sterile filtered at 0.2 μm. The filtrate was used directly for lipase enzyme activity characterization without further treatment. Activity was measured in tributyrin / agarose emulsion, for example, as described in Example 2 below or by the IDF method. In particular, the IDF method was used for the administration of Examples 3 to 6.

[0084] Example 2 - Enzyme activity towards triacylglycerol (TAG) substrates In a 96-well plate assay, short-chain fatty acids (C4, substrate tributyrin) and long-chain fatty acids (C 16 and C 18 The enzymatic activity of each microbial lipase (SEQ ID NOs: 1 to 11) disclosed herein against short-chain fatty acids (C4, substrate tributyrin) and long-chain fatty acids (C6, substrate olive oil) was measured as follows in a 96-well plate assay. 16 and C 18The same procedure was also performed for a commercially available microbial lipase from Rhizomucor miehei (Palatase®, Palatase® 20000 L, ≥ 20,000 U / g, available from Novozymes A / S) to measure the enzyme activity against the substrate olive oil. The commercially available microbial lipase from Rhizomucor miehei is referred to herein as commercial microbial lipase A.

[0085] Enzyme activity toward short-chain fatty acids in plate culture: Tributyrin / agarose emulsions were prepared by dissolving 1.4% low-melting-point agarose in 250 mM sodium acetate buffer, pH 5.25, containing 40 mM CaCl2, adding 1% (v / v) tributyrin to the solubilized agarose at 60°C, and then emulsifying for 2 minutes using an IKA T25 Ultra-Turrax emulsifier at speed 13.5. A total volume of 50 μL of emulsion was transferred to each well of a clear 96-well assay plate and allowed to solidify at room temperature. The lipase reaction was initiated by overlaying 20 μL of enzyme onto the emulsion, where 20 μL of enzyme corresponded to 20 μL of filtrate obtained from Example 1. Following clearance of emulsion turbidity due to enzymatic hydrolysis of the substrate tributyrin, absorbance was measured at 600 nm using a plate reader. Each assay was calibrated using various dilutions of a commercially available microbial lipase A with known enzymatic activity.

[0086] Enzyme activity toward long-chain fatty acids in plate culture: Olive oil / agarose emulsions were prepared by dissolving 1.4% low-melting-point agarose in 250 mM sodium acetate buffer, pH 5.25, containing 40 mM CaCl2, adding 2.5% (v / v) olive oil and 0.001% rhodamine B (w / v) to the solubilized agarose at 60°C, and then emulsifying for 2 minutes using an IKA T25 Ultra-Turrax emulsifier at speed 13.5. A total volume of 150 μL of emulsion was transferred to each well of a non-transparent black 96-well assay plate and allowed to solidify at room temperature. The lipase reaction was initiated by layering 30 μL of enzyme onto the emulsion, where 30 μL of enzyme corresponded to 30 μL of filtrate from Example 1. Following the binding of long-chain fatty acids released by enzymatic hydrolysis of the substrate olive oil to rhodamine B, fluorescence was measured in a plate reader at excitation of 355 nm and emission of 590 nm. Each assay was calibrated using various dilutions of a commercially available microbial lipase A with known enzymatic activity.

[0087] Short chain fatty acid specificity C4 / C 16 ~C 18 The substrates tributyrin (C4) and olive oil (C 16 ~C 18 The hydrolytic activity was determined for each microbial lipase disclosed herein by calculating the ratio of the hydrolytic activity to the total hydrolytic activity of the microbial lipase.

[0088] The ratio C4 / C of each lipase (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, and / or SEQ ID NO: 11) 16 ~C 18 The ratio C4 / C of the commercially available lipase (A) of the microorganism used 16 ~C 18Table 1 shows that the microbial lipases disclosed herein (SEQ ID NOs: 1 to 11) have higher specificity for the short-chain fatty acid butyric acid (C4) when compared to commercially available microbial lipase A, and in particular, the microbial lipases disclosed herein (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 and / or SEQ ID NO: 11) have 2 to 770 times higher C4-fatty acid specificity than commercially available microbial lipase A.

[0089] Table 1. Short-chain fatty acid specificity (C4 / C6) of lipases having 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 and commercial microbial lipase A, and their respective improvement factors (IF4) determined using the TAG substrate tributyrin and olive oil. 16 ~C 18 ).

[0090] [Table 1]

[0091] Example 3 - Enzyme activity in cream The lipases disclosed herein (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, or SEQ ID NO:11) were analyzed for their enzymatic activity on milk fat and the free fatty acids released were quantified (Table 2). The same procedure was performed for a commercially available microbial lipase (commercial microbial lipase A) from Rhizomucour miehei (Palatase®) available from Novozymes A / S.

[0092] Lipase activity against milk fat in cream was measured as follows: Fresh whipping cream (38% lipid) and an equal volume of water were mixed by stirring for 30 minutes. The enzymatic reaction was initiated by adding 20 μL of lipase (from a 4000 LFU / L stock solution) to 1 mL of the cream / water mixture and continued for 20 hours at 37°C. This enzyme dose specifically applies to both commercial microbial lipase A and lipases of SEQ ID NOS: 1-11. Subsequently, free fatty acids were extracted from the reaction mixture and quantified by gas chromatography-mass spectrometry (GC-MS) using standard protocols for fatty acid quantification. Alternatively, the disclosure of Jong C., de and Badings HT in J. High Resolution Chromatography. 13, 84-98 (1990) can also be used.

[0093] The resulting fatty acid amounts of the lipase reaction were corrected against the respective amounts measured for similar cream samples without added lipase.

[0094] Subsequently, the effect of butyric acid (C) was compared with that of commercially available microbial lipase A. 4:0 ), caproic acid (C 6:0 ), caprylic acid (C 8:0 ), capric acid (C 10:0 ), and the specificity of each improvement factor were determined based on data obtained from lipolysis in cream (Table 2). Short-chain fatty acid specificity (C4 / C 16 ~C 18 ;C6 / C 16 ~C 18 ) to C 4:0 or C 6:0 Molar fraction of and C 16:0 , C 18:0 , C 18:1 , and C 18:2 The improvement factors IF4 and IF6 were calculated by the ratio of the molar fractions of C 4:0 and C 6:0 The specificity values ​​of the enzymes were calculated by dividing them by the respective values ​​of the commercial lipase A. Medium-chain fatty acid specificity (C8 / C16 ~C 18 ;C 10 / C 16 ~C 18 ) to C 8:0 or C 10:0 Molar fraction of and C 16:0 , C 18:0 , C 18:1 , and C 18:2 The improvement factors IF8 and IF10 were calculated by the ratio of the molar fractions of C 8:0 and C 10:0 The specificity values ​​of the enzymes were calculated by dividing them by the respective values ​​of the commercial lipase A.

[0095] C4 / C of each lipolytic lipase 16 ~C 18 The ratio is the C of each lipolytic lipase. 4:0 C 16:0 , C 18:0 , C 18:1 , and C 18:2 The ratio C4 / C was determined by dividing by the sum of 16 ~C 18 is the chain length C 16 ~C 18 This suggests the specificity of each lipase for the release of fatty acids with C4, C5, and C6 from milk fat.

[0096] C6 / C of each lipolytic lipase 16 ~C 18 The ratio is the C of each lipolytic lipase. 6:0 C 16:0 , C 18:0 , C 18:1 , and C 18:2 The ratio C6 / C 16 ~C 18 is the chain length C 16 ~C 18 This suggests the specificity of each lipase for the release of fatty acids with caproic acid (C6) from milk fat, relative to the release of fatty acids with caproic acid (C6) from milk fat.

[0097] C8 / C of each lipolytic lipase 16 ~C 18The ratio is the C of each lipolytic lipase. 8:0 C 16:0 , C 18:0 , C 18:1 , and C 18:2 The ratio C8 / C 16 ~C 18 is the chain length C 16 ~C 18 This suggests the specificity of each lipase for the release of fatty acids with a C8 group, whereas the release of caprylic acid (C8) from milk fat is different.

[0098] C of each lipolytic lipase 10 / C 16 ~C 18 The ratio is the C of each lipolytic lipase. 10:0 C 16:0 , C 18:0 , C 18:1 , and C 18:2 The ratio C was determined by dividing by the sum of 10 / C 16 ~C 18 is the chain length C 16 ~C 18 The effect of capric acid (C) from milk fat on the release of fatty acids with 10 This suggests the specificity of each lipase for the release of .

[0099] Table 2. Molar fraction of fatty acids released in cream by commercial microbial lipase A and 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 and SEQ ID NO:11

[0100] [Table 2]

[0101] Table 3. Short- and medium-chain fatty acid specificity (C4 / C6) in cream by commercially available microbial lipase A and 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, and SEQ ID NO: 11 16~C 18 , C6 / C 16 ~C 18 , C8 / C 16 ~C 18 , C 10 / C 16 ~C 18 ) and improvement factors (IF4, IF6, IF8, IF10)

[0102] [Table 3]

[0103] All of the microbial lipolytic enzymes disclosed herein (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 and / or SEQ ID NO: 11) exhibit an IF4 higher than 1 in cream, indicating that they are able to selectively degrade the short chain fatty acid butyrate (C) compared to commercial lipase A. 4:0 ) (Table 3), and therefore can be used in combination with the short-chain fatty acid butyric acid (C ) to, for example, reduce the soapiness of food products and enhance the butyric acid flavor. 4:0 ) is necessary and can be successfully used in the process of producing the desired food product.

[0104] The microbial lipolytic enzymes having SEQ ID NOs: 1 to 7 and SEQ ID NOs: 9 to 11 showed an IF6 higher than 1 in cream, and they were able to degrade the short-chain fatty acid caproic acid (C ) compared to commercial lipase A. 6:0 ) means that it has higher specificity for

[0105] The microbial lipolytic enzymes having SEQ ID NOs: 1 to 6 and SEQ ID NOs: 8 to 11 showed an IF8 higher than 1 in cream, and they were found to be more effective in the degradation of the medium-chain fatty acid caprylic acid (C ) compared to commercial lipase A. 8:0 ) means that it has higher specificity for

[0106] The microbial lipolytic enzymes having SEQ ID NOs: 1 to 3, 6 to 8, and 10 to 11 showed an IF10 higher than 1 in cream, and they were able to degrade the medium-chain fatty acid capric acid (C ) compared to commercial lipase A. 10:0 ) means that it has higher specificity for

[0107] The microbial lipolytic enzyme having SEQ ID NO: 1 was found to produce a higher C fatty acid content than other fatty acids in cream. 4:0 The same applies to the microbial lipolytic enzyme having SEQ ID NO: 3 or SEQ ID NO: 9 (Table 2).

[0108] Example 4 - Enzyme activity in cheese SEQ ID NO: 1 was further tested in cheese, specifically Feta and Provolone cheese (Table 4). Example 4 was carried out using the same dosage of enzyme (commercial microbial lipase A and SEQ ID NO: 1), specifically a dosage of about 1.7 LFU / L of enzyme per liter of milk was used for the production of Feta cheese and 5-10x for Provolone cheese. Doses were determined by the IDF method.

[0109] Microbial lipase SEQ ID NO: 1 was added during the cheese-making process. The cheese-making process, for example, of Feta or Provolone cheese, is well known in the art and to those skilled in the art. Subsequently, free fatty acids were extracted from the cheese after one month of aging and quantified by GC-MS or as disclosed in Example 3. The fatty acid amounts of the resulting lipase reaction were corrected against the respective amounts measured for similar cheeses without the addition of lipase. Short-chain fatty acid specificity was determined by the C 4:0 or C 6:0 and the mole fraction of C 16:0 , C 18:0 , C 18:1 , and C 18:2 The improvement factors IF4 and IF6 were calculated by the ratio of the molar fraction of C 4:0 or C 6:0 The specificity values ​​of the enzymes were calculated by dividing them by the respective values ​​of the commercial lipase A.

[0110] Table 4. Molar fractions of fatty acids released in Feta and Provolone cheeses by commercial microbial lipase A and SEQ ID NO: 1 after 1 month of aging, as well as short-chain fatty acid specificity (C4 / C 16 ~C 18 and C6 / C 16 ~C 18 ) and improvement factors (IF4 and IF6)

[0111] [Table 4]

[0112] SEQ ID NO: 1 exhibits superior C activity compared to commercially available microbial lipase A in feta and provolone cheeses. 4:0 -has a higher specificity for fatty acids and C compared to other fatty acids in both Feta and Provolone cheeses 4:0 -fatty acids, which is consistent with the results obtained with cream (Example 3). Furthermore, SEQ ID NO: 1 also exhibited a higher specificity for C-fatty acids than commercial microbial lipase A in feta and provolone cheeses. 6:0 - also has a higher specificity for fatty acids, which is also in agreement with the results obtained with cream (Example 3).

[0113] SEQ ID NO: 1 also demonstrated a C activity when compared to a commercial microbial lipase A in feta cheese. 8:0 , C 10:0 , C 14:0 , C 16:0 , C 18:0 , C 18:1 or C 18:2 Furthermore, SEQ ID NO: 1 also exhibits lower specificity for medium to long chain fatty acids such as C when compared to commercial microbial lipase A in Provolone cheese. 8:0 , C 10:0 , C 12:0 , C 14:0 , C 16:0 , C 18:0 , C 18:1 or C 18:2These results are consistent with those obtained with cream (Example 3). Thus, cheeses produced using a microbial lipase comprising SEQ ID NO: 1, or where the microbial lipase is the microbial lipase of SEQ ID NO: 1, have a weaker soapy flavor and a stronger butyric flavor than cheeses produced with known microbial lipases. The same results are expected for 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, and / or SEQ ID NO: 11.

[0114] Identical results are also expected for other types of cheese, such as Pecorino, Parmesan, Grana Padano, Parmigiano-Reggiano, Romano, Chester, Dumbo, Manchego, Saint Paulin, Cheddar, Monterey, Colby, Edam, Gouda, Muenster, Swiss-type, Gruyère, and Emmental; pasta filata cheeses such as Mozzarella and Queso Fresco; fresh cheeses such as Ricotta, Cream Cheese, Neuchâtel, or Cottage Cheese; cream cheese, or white-mold cheeses such as Brie and Camembert, or Gorgonzola and Danish Blue Cheese; processed cheese, cheese-like products, or blue-mold cheeses such as enzyme-treated cheeses.

[0115] The microbial lipases disclosed herein are responsible for enhanced C4-fatty acid specificity compared to prior art microbial lipases, which leads to reduced soapiness in cheese and enhanced butyric flavor in cheese.

[0116] Example 5 - Sensory evaluation Sensory evaluation of the cheeses, especially the 3-month aged Feta cheese, was also carried out by rating the flavor according to the sensory parameters of butyric and soapy notes on a scale from 1 (none) to 6 (strong). The Feta cheeses were produced using the same conditions using Commercial A, SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5. The amount or dosage of the enzyme (commercial microbial lipase A, SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5) used was 1.7 LFU / L milk as determined by the IDF method (Table 5).

[0117] Table 5. Sensory evaluation performed on Feta cheese aged for 3 months using commercial microbial lipases A, SEQ ID NO: 1, SEQ ID NO: 3 and SEQ ID NO: 5

[0118] [Table 5]

[0119] Table 5 shows the reduction in soapiness in 3 month aged Feta cheese when SEQ ID NO: 1, 3 or 5 was used compared to commercial Lipase A, and at the same time an increase in butyric acid flavor was observed in the same samples.

[0120] Example 6 - Enzyme activity in cheese SEQ ID NOs: 1, 2, 3, 6, 8, 10 and 11 were further tested in cheese, specifically Feta cheese. Example 6 was performed using the same dosage of enzymes (commercial microbial lipase A and SEQ ID NOs: 1, 2, 3, 6, 8, 10 and 11), specifically the enzyme dosage was about 2.3 LFU / L milk, as determined by the IDF method, for the production of Feta cheese.

[0121] Microbial lipases SEQ ID NOs: 1, 2, 3, 6, 8, 10, and 11 were added individually during the cheese-making process. Free fatty acids were subsequently extracted from the cheese after 1 month (Table 6) and 2 months (Table 8) of ripening and quantified by GC-MS or as disclosed in Example 3. The fatty acid amounts of the resulting lipase reactions were corrected to the respective amounts measured for similar cheeses without added lipase. Short-medium chain fatty acid specificity was determined by the C 4:0 , C 6:0 , C 8:0 or C 10:0 and the mole fraction of C 16:0 , C 16:1 , C 18:0 , C 18:1 , C 18:2 , and C 18 The improvement factors IF4, IF6, IF8 and IF10 were calculated by the ratio of the molar fraction of C to the sum of the molar fractions of C:3 (Tables 7 and 9). 4:0 , C 6:0 , C 8:0 or C 10:0 The specificity values ​​of the enzymes were calculated by dividing the specificity values ​​of the enzymes by the respective values ​​of the commercial lipase A (Tables 7 and 9).

[0122] Table 6. Molar fraction of fatty acids released in Feta cheese by commercial microbial lipase A and SEQ ID NOs: 1, 2, 3, 6, 10 and 11 after 1 month of aging. All lipases were dosed at 2.3 LFU / L.

[0123] [Table 6]

[0124] Table 7. Short-medium chain fatty acid specificity (C4 / C6) in Feta cheese by commercial microbial lipase A and SEQ ID NOs: 1, 2, 3, 6, 10 and 11 after 1 month of aging 16 ~C 18 , C6 / C 16 ~C 18 , C8 / C 16 ~C 18 , C 10 / C 16 ~C 18) and improvement factors (IF4, IF6, IF8, IF10). All lipases were administered at 2.3 LFU / L.

[0125] [Table 7]

[0126] All tested microbial lipolytic enzymes show an IF4 higher than 1 in Feta cheese after 1 month of ripening. Thus, the tested enzymes have a higher affinity for the short chain fatty acid butyrate (C) compared to commercial lipase A. 4:0 ) (Table 7), and therefore can be used in combination with the short-chain fatty acid butyric acid (C ), for example, to reduce soapiness and enhance butyric flavor in food products. 4:0 ) is necessary and can be successfully used in the process of producing the desired food product. This observation of an IF higher than 1 for the tested lipase is consistent with the results of Example 3, where data was obtained with cream. Moreover, specifically for SEQ ID NO: 1, this observation is also consistent with the results of Example 4.

[0127] Furthermore, these enzymes showed IF6, IF8 and IF10 higher than 1 in Feta cheese after 1 month of ripening, and they showed a higher IF6, IF8 and IF10 than commercial lipase A, except for SEQ ID NO: 2, which had an IF10 lower than 1, for the short chain fatty acid caproic acid (C 6:0 ) and medium-chain fatty acid caprylic acid (C 8:0 ) and capric acid (C 10:0 ) and the results obtained for these sequences (SEQ ID NOs: 1, 2, 3, 6, 10 and 11) and C 6:0 -Fatty acids, C 8:0 -Fatty acids and C 10:0 Their specificity for fatty acids is substantially consistent with the results obtained in cream (Example 3).

[0128] In conclusion, after one month of aging, the results obtained in cheese (Examples 4 and 6) are consistent with those obtained in cream (Example 3), and show that the results obtained in cream can be used to plausibly extrapolate the behavior of untested sequences from cream to cheese.

[0129] Table 8. Molar fraction of fatty acids released in Feta cheese by commercial microbial lipase A and SEQ ID NOs: 1, 2, 3, 6, 8, 10 and 11 after 2 months of ripening. All lipases were dosed at 2.3 LFU / L.

[0130] [Table 8]

[0131] Table 9. Short-medium chain fatty acid specificity (C4 / C6) in Feta cheese by commercial microbial lipase A and SEQ ID NOs: 1, 2, 3, 6, 8, 10 and 11 after 2 months of aging 16 ~C 18 , C6 / C 16 ~C 18 , C8 / C 16 ~C 18 , C 10 / C 16 ~C 18 ) and improvement factors (IF4, IF6, IF8, IF10). All lipases were administered at 2.3 LFU / L.

[0132] [Table 9]

[0133] All tested microbial lipolytic enzymes show an IF4 higher than 1 in Feta cheese after 2 months of ripening. Thus, the tested enzymes are better able to release the short chain fatty acid butyrate (C) compared to commercial lipase A. 4:0 ) (Table 7), and therefore can be used in combination with the short-chain fatty acid butyric acid (C ), for example, to reduce soapiness and enhance butyric flavor in food products. 4:0) is necessary and can be successfully used in the process of producing the desired food product.

[0134] The microbial lipolytic enzymes having SEQ ID NOs: 1, 2, 3, 6, 8, 10 and 11 exhibited higher IF6 and IF8 than commercial lipase A, indicating that they were able to degrade the short-chain fatty acid caproic acid (C 6:0 ) and medium-chain fatty acid caprylic acid (C 8:0 ) means that it has higher specificity for

[0135] With the exception of SEQ ID NO: 3, the tested enzymes also showed an IF10 higher than 1, and therefore, a higher IF10 of the medium chain fatty acid capric acid (C 10:0 ) has higher specificity. Additionally, and as previously mentioned, the present invention relates to flavor enhancement, but also involves shortening the aging period of aged cheese. A comparison between the free fatty acid profiles after one or two months of aging shows that it is possible to successfully shorten the aging period from (more than) two months to (less than) one month while still providing cheese with the desired short chain fatty acid profile.

[0136] Identical results to those disclosed in Example 6 are also expected for other types of cheese, such as Pecorino, Parmesan, Grana Padano, Parmigiano-Reggiano, Romano, Chester, Dumbo, Manchego, Saint Paulin, Cheddar, Monterey, Colby, Edam, Gouda, Muenster, Swiss-type, Gruyere, Emmental; pasta filata cheeses such as mozzarella or queso fresco; fresh cheeses such as ricotta, cream cheese, Neuchâtel, or cottage cheese; cream cheese, or white-mold cheeses such as Brie or Camembert, or blue-mold cheeses such as Gorgonzola or Danish blue cheese; processed cheese, cheese-like products, or enzyme-treated cheeses.

[0137] In conclusion, the present invention discloses a microbial wild-type lipase that has a higher specificity for short-chain fatty acids, particularly C4-fatty acids, compared to known prior art microbial lipases, leading to reduced soapiness and enhanced butyric flavor in food products such as cheese. Therefore, the present invention provides a microbial wild-type lipase that satisfies vegetarian and / or kosher requirements while simultaneously exhibiting a higher specificity for short-chain fatty acids, particularly C4-fatty acids, from milk fat and / or other lipids, and a lower specificity for medium- to long-chain fatty acids, particularly medium- to long-chain fatty acids from milk fat and / or other lipids, thereby avoiding the very unpleasant soapy flavor in cheese-like food products.

[0138] The present invention is applicable to other types of food products, the purpose of which is to reduce the unpleasant soapy taste of said food products. Finally, if necessary, the ripening period of food products, particularly Feta cheese, can be reduced from two months to one month while still maintaining an appropriate and desired short chain fatty acid profile.

[0139] In the context of describing the present invention (particularly in the claims that follow), the use of the terms "a," "an," "the," and similar referents should be construed to cover both the singular and the plural unless otherwise stated or clearly contradicted by context. The terms "comprise," "have," "including," and "containing" should be construed as open-ended terms (i.e., meaning "including, but not limited to") unless otherwise noted. The recitation of ranges of values ​​herein is intended to serve as a shorthand method of individually referring to each individual value falling within the range, unless otherwise indicated herein, and each individual value is incorporated herein as if it were individually listed herein. All methods described herein can be performed in any suitable order unless otherwise indicated herein or clearly contradicted by context. The use of any and all examples or exemplary language (e.g., "etc.") provided herein is intended merely to better clarify the invention and does not limit the scope of the invention unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element essential to the practice of the invention.

[0140] A list of preferred embodiments in claim form

[0141] 1. has at least 90% identity to 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 or SEQ ID NO:11, preferably SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, more preferably SEQ ID NO:1, SEQ ID NO:3 or SEQ ID NO:11, even more preferably SEQ ID NO:1, or having at least 95%, 96%, 97%, 98%, 99% identity to 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 or SEQ ID NO:11, preferably SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, more preferably SEQ ID NO:1, SEQ ID NO:3 or SEQ ID NO:11, even more preferably SEQ ID NO:1, containing an amino acid sequence, or wherein the sequence is 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 or SEQ ID NO: 11, preferably SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 9 or SEQ ID NO: 11, more preferably SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 11, even more preferably SEQ ID NO: 1, Lipase.

[0142] 2. The lipase is C 10 2. The lipase according to item 1, which has a higher specificity for the release of C4-fatty acids from milk fat and / or dairy compositions containing other fats when compared to the release of C4-fatty acids.

[0143] 3. The lipase releases C6-fatty acids compared to C8-fatty acids. 12 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 18:0 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 16 -When compared to the release of fatty acids, or C8-fatty acids ~ C 18:2 -When compared to the release of fatty acids, C 10 -Fatty acid~C 18:1 -When compared to the release of fatty acids, C 12 -Fatty acid~C 18:0 -When compared to the release of fatty acids, C 14 -Fatty acid~C 16:0-relative to the release of fatty acids, or C 16 ~C 18 3. The lipase according to any one of items 1 to 2, which has a higher specificity for the release of C4-fatty acids from milk fat and / or dairy compositions comprising other fats compared to the release of C4-fatty acids.

[0144] 4. The lipase is preferably at a pH of less than 7, preferably at a pH of 6.6 to 6.8, less than pH 6, preferably at a pH of 3.8 to 5.6, more preferably at a pH of 4.4 to 5.4, and even more preferably at a pH of 4.6 to 5.2, 10 - compared to the release of fatty acids, or compared to the release of C6-fatty acids, or compared to the release of C8-fatty acids, 12 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 16 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 18:3 -relative to the release of fatty acids, or C 16 ~C 18 4. The lipase according to any one of items 1 to 3, which has a higher specificity for the release of C4-fatty acids from milk fat and / or dairy compositions comprising other fats compared to the release of C4-fatty acids.

[0145] 5. The lipase is reacted at a temperature of less than 20°C, preferably less than 15°C, more preferably less than 10°C, and even more preferably between 5 and 8°C. 10 - compared to the release of fatty acids, or compared to the release of C6-fatty acids, or compared to the release of C8-fatty acids, 12 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 16 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 18:3 -relative to the release of fatty acids, or C 16~C 18 5. The lipase according to any one of items 1 to 4, which has a higher specificity for the release of C4-fatty acids from milk fat and / or dairy compositions comprising other fats compared to the release of C4-fatty acids.

[0146] 6. 6. The lipase according to any one of items 1 to 5, wherein the lipase is a microbial lipase, preferably an isolated microbial lipase, a recombinant microbial lipase, or a synthetic microbial lipase.

[0147] 7. 7. An isolated, recombinant, or synthetic DNA sequence encoding the lipase according to any one of Items 1 to 6, wherein the isolated, recombinant, or synthetic DNA sequence is selected from sequences having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, or SEQ ID NO: 23.

[0148] 8. 8. The DNA sequence according to any one of items 1 to 7, further comprising a signal peptide sequence.

[0149] 9. A vector comprising the isolated, recombinant or synthetic DNA sequence according to any one of items 7 to 8.

[0150] 10. A host cell, preferably a genetically modified host cell, comprising any of the sequences according to any one of items 1 to 8 or the vector according to item 9.

[0151] 11. 11. The host cell according to any one of items 1 to 10, wherein the host cell is selected from Pichia pastoris, Aspergillus or Bacillus subtilis, preferably Pichia pastoris or B. subtilis, more preferably Pichia pastoris.

[0152] 12. 1. A method for preparing a food product, comprising the steps of: or using a lipase comprising an amino acid sequence which has at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to 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 or SEQ ID NO:11, preferably having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, or using a DNA sequence encoding a lipase; wherein the DNA sequence is selected from sequences having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, or SEQ ID NO:23, preferably having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:14, SEQ ID NO:20, SEQ ID NO:22, or SEQ ID NO:23; preferably, the DNA sequence is an isolated, recombinant, or synthetic DNA sequence, more preferably wherein the DNA sequence comprises a signal peptide sequence.

[0153] 13. The lipase is C 10- compared to the release of C8-fatty acids, compared to the release of C6-fatty acids, 12 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 18:0 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 16 -When compared to the release of fatty acids, or C8-fatty acids ~ C 18:2 -When compared to the release of fatty acids, C 10 -Fatty acid~C 18:1 -When compared to the release of fatty acids, C 12 -Fatty acid~C 18:0 -When compared to the release of fatty acids, C 14 -Fatty acid~C 16:0 -relative to the release of fatty acids, or C 16 ~C 18 13. The method according to claim 12, wherein the method has a higher specificity for the release of C4-fatty acids from milk fat and / or other fat-containing dairy compositions when compared to the release of C4-fatty acids.

[0154] 14. The lipase is preferably at a pH of less than 7, preferably at a pH of 6.6 to 6.8, less than pH 6, preferably at a pH of 3.8 to 5.6, more preferably at a pH of 4.4 to 5.4, and even more preferably at a pH of 4.6 to 5.2, 10 - compared to the release of fatty acids, or compared to the release of C6-fatty acids, or compared to the release of C8-fatty acids, 12 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 16 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 18:3 -relative to the release of fatty acids, or C 16 ~C 1814. The method according to any one of items 12 to 13, having a higher specificity for the release of C4-fatty acids from milk fat and / or dairy compositions comprising other fats when compared to the release of C4-fatty acids.

[0155] 15. The lipase is reacted at a temperature of less than 20°C, preferably less than 15°C, more preferably less than 10°C, and even more preferably between 5 and 8°C. 10 - compared to the release of fatty acids, or compared to the release of C6-fatty acids, or compared to the release of C8-fatty acids, 12 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 16 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 18:3 -relative to the release of fatty acids, or C 16 ~C 18 15. The method according to any one of items 12 to 14, having a higher specificity for the release of C4-fatty acids from milk fat and / or dairy compositions comprising other fats when compared to the release of fatty acids.

[0156] 16. 16. The method according to any one of Items 12 to 15, wherein the lipase is a microbial lipase, preferably an isolated microbial lipase, a recombinant microbial lipase, or a synthetic microbial lipase.

[0157] 17. 17. The method according to any one of items 12 to 16, wherein the food product is a dairy product selected from cheese, processed cheese, cheese-like products, enzyme-treated cheese, butter, yogurt, cream, or seasonings.

[0158] 18. 18. The method according to any one of items 12 to 17, wherein the dairy product is cheese, preferably wherein the cheese is feta cheese, provolone cheese, pecorino cheese, parmesan cheese, grana padano cheese, parmigiano reggiano cheese, romano cheese, chester cheese, dambo cheese, manchego cheese, saint paulin cheese, cheddar cheese, monterey cheese, colby cheese, edam cheese, gouda cheese, muenster cheese, swiss type cheese, gruyere cheese or emmental cheese; or pasta filata cheese such as mozzarella or queso fresco cheese; or fresh cheese such as ricotta, cream cheese, neuchâtel or cottage cheese; or cream cheese, or white mould cheese such as brie or camembert cheese, or blue mould cheese such as gorgonzola or danish blue cheese.

[0159] 19. The food product is a dairy alternative product, preferably a dairy alternative cheese or a vegan cheese, preferably wherein the lipase is selected from the group consisting of C 10 - compared to the release of C8-fatty acids, compared to the release of C6-fatty acids, 12 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 18:0 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 16 -When compared to the release of fatty acids, or C8-fatty acids ~ C 18:2 -When compared to the release of fatty acids, C 10 -Fatty acid~C 18:1 -When compared to the release of fatty acids, C 12 -Fatty acid~C 18:0 -When compared to the release of fatty acids, C 14 -Fatty acid~C 16:0 -relative to the release of fatty acids, or C 16 ~C 18- the method according to item 12, having a higher specificity for the release of C4-fatty acids from lipid-containing compositions such as vegetable oils or water-in-oil emulsions when compared to the release of fatty acids.

[0160] 20. 1. A food product comprising a lipase, wherein the lipase comprises an amino acid sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to 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 or SEQ ID NO:11, preferably having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11; and wherein the food product is C 10 -fatty acids, or C6-fatty acids, C8-fatty acids, C 12 -Fatty acids, C 14 -Fatty acids, C 16 -Fatty acids, C 18:1 -Fatty acids, C 18:2 -fatty acid or C 18:3 -contains more C4-fatty acids than other fatty acids selected from the fatty acids; and / or wherein the food product is C 10 -fatty acids, or C8-fatty acids, C 12 -Fatty acids, C 14 -Fatty acids, C 16 -Fatty acids, C 18:1 -Fatty acids, C 18:2 -fatty acid or C 18:3 - A food product containing a higher amount of C6-fatty acids compared to other fatty acids selected from fatty acids.

[0161] twenty one. the food product is cheese, processed cheese, cheese-like product, or a dairy product selected from butter, yogurt, cream, seasonings, preferably wherein the cheese is feta cheese, provolone cheese, pecorino cheese, parmesan cheese, grana padano cheese, parmigiano reggiano cheese, romano cheese, chester cheese, dambo cheese, manchego cheese, saint paulin cheese, cheddar cheese, monterey cheese, colby cheese, edam cheese, gouda cheese, muenster cheese, swiss type cheese, gruyere cheese, or emmental cheese; or pasta filata cheese such as mozzarella or queso fresco cheese; or fresh cheese such as ricotta, cream cheese, neuchâtel or cottage cheese; or cream cheese, or white mould cheese such as brie or camembert cheese, or blue mould cheese such as gorgonzola or danish blue cheese, more preferably feta cheese or provolone cheese; 21. The food product according to item 20, wherein the food product is a dairy alternative product such as a milk alternative cheese or a vegan cheese.

[0162] twenty two. 1. Use of a lipase for producing a food product, wherein the food product is a dairy product selected from cheese, processed cheese, enzyme-modified cheese (EMC), cheese-like products, butter, yogurt, cream, or seasonings, preferably feta cheese, provolone cheese, pecorino cheese, parmesan cheese, grana padano cheese, parmigiano reggiano cheese, romano cheese, chester cheese, dambo cheese, manchego cheese, saint paulin cheese, cheddar cheese, monterey cheese, colby cheese, edam cheese. or pasta filata cheeses such as mozzarella or queso fresco; or fresh cheeses such as ricotta, cream cheese, Neuchâtel or cottage cheese; or cream cheese, or white-veined cheeses such as brie or camembert, or blue-veined cheeses such as gorgonzola or danish blue cheese; more preferably the cheese is feta or provolone, wherein the food product is a dairy alternative product, such as a dairy alternative cheese or a vegan cheese; and Use of a lipase wherein the lipase comprises an amino acid sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to 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 or SEQ ID NO:11, preferably having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11.

[0163] twenty three. More C8-fatty acids, C 10 -Fatty acids, C 12 -Fatty acids, C 14 -Fatty acids, C 16 -Fatty acids, C18:0 -Fatty acids, C 18:1 -Fatty acids, C 18:2 -fatty acids, or C 18:3 10. Use of a lipase to produce or release more C4- and / or C6-fatty acids from a dairy composition comprising milk fat and / or other lipids, such as vegetable oils, compared to producing or releasing fatty acids, wherein the lipase comprises an amino acid sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to 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 or SEQ ID NO:11, preferably having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, and preferably wherein the lipase is selected from the group consisting of C 10 - compared to the release of C8-fatty acids, compared to the release of C6-fatty acids, 12 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 18:0 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 16 -When compared to the release of fatty acids, or C8-fatty acids ~ C 18:2 -When compared to the release of fatty acids, C 10 -Fatty acid~C 18:1 -When compared to the release of fatty acids, C 12 -Fatty acid~C 18:0 -When compared to the release of fatty acids, C 14 -Fatty acid~C 16:0 -relative to the release of fatty acids, or C 16 ~C 18 - Use of lipases that have a higher specificity for the release of C4-fatty acids from milk fat and / or dairy compositions containing other fats when compared to the release of fatty acids.

[0164] twenty four. More C8-fatty acids in food products, C 10 -Fatty acids, C 12 -Fatty acids, C 14 -Fatty acids, C 16 -Fatty acids, C 18:0 -Fatty acids, C 18:1 -Fatty acids, C 18:2 -fatty acids, or C 18:3 - use of a lipase to produce or release more C4- and / or C6-fatty acids in a food product compared to producing or releasing fatty acids, wherein the lipase comprises an amino acid sequence that has at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to 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 or SEQ ID NO:11, preferably at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11, Preferably, wherein the food product is a dairy product selected from cheese, processed cheese, enzyme-treated cheese, cheese-like products, butter, yogurt, cream, or seasonings; or Preferably, the use of lipase wherein the food product is a dairy alternative product such as a milk alternative cheese or a vegan cheese.

[0165] twenty five. 25. Use according to item 24, wherein the food product is a dairy product selected from cheese, and wherein the cheese is Feta cheese, Provolone cheese, Pecorino cheese, Parmesan cheese, Grana Padano cheese, Parmigiano-Reggiano cheese, Romano cheese, Chester cheese, Dumbo cheese, Manchego cheese, Saint Paulin cheese, Cheddar cheese, Monterey cheese, Colby cheese, Edam cheese, Gouda cheese, Muenster cheese, Swiss-type cheese, Gruyère cheese, or Emmental cheese; or Pasta filata cheese such as Mozzarella or Queso Fresco; or Fresh cheese such as Ricotta, Cream cheese, Neuchâtel or Cottage cheese; or Cream cheese, or White mould cheese such as Brie or Camembert, or Blue mould cheese such as Gorgonzola or Danish Blue cheese, preferably Feta cheese or Provolone cheese.

[0166] 26. a lipase consisting of an amino acid sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to 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 or SEQ ID NO:11, preferably having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:9 or SEQ ID NO:11; wherein the lipase is expressed in Pichia pastoris and 10 - compared to the release of C8-fatty acids, compared to the release of C6-fatty acids, 12 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 18:0 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C16 -When compared to the release of fatty acids, or C8-fatty acids ~ C 18:2 -When compared to the release of fatty acids, C 10 -Fatty acid~C 18:1 -When compared to the release of fatty acids, C 12 -Fatty acid~C 18:0 -When compared to the release of fatty acids, C 14 -Fatty acid~C 16:0 -relative to the release of fatty acids, or C 16 ~C 18 - a lipase that has a higher specificity for the release of C4-fatty acids from milk fat and / or dairy compositions containing other fats when compared to the release of fatty acids.

[0167] 27. 27. The lipase according to item 26, wherein the lipase is used in any of the methods or uses disclosed herein.

[0168] References Non-patent literature Iwai, M. et al. Studies on Lipase II. Hydrolytic and esterifying actions of crystalline lipase of Aspergillus niger. J. Gen. Appl. Microbiol. 10, 13-22 (1964) Oi, S. et al. Purification and Some Properties of a Fungal Lipase Preparation Used for Milk Flavouring. Agr. Biol. Chem. 33, 729-38 (1969) Somkuti, GA et al. Lipase Activity of Mucor pusillus. Appl. Microbiol. 16, 617-19 (1968) Kamoun, J. et al. Biochemical characterization of Yarrowia lipolytica LIP8, a secreted lipase with a cleavable C-terminal region. Biochim. Biophys. Acta 1851, 129-140 (2015) Aloulou, A. et al. Purification and biochemical characterization of the LIP2 lipase from Yarrowia lipolytica. Biochim. Biophys. Acta - Mol. Cell Biol. Lipids 1771, 228-237 (2007) Sheng, J., Wang, F., Wang, H. & Sun, M. Cloning, characterization and expression of a novel lipase gene from marine psychrotrophic Yarrowia lipolytica. Ann. Microbiol. 62, 1071-1077 (2012) J. Sambrook, E. F. Fritsch, and T. Maniatis, 1989, Molecular Cloning: A Laboratory manual, 2 nd edition, books 1-3, Cold Spring Harbor Laboratory Press Jong C., de and Badings H.T. Determination of free fatty acids in milk and cheese procedures for extraction, clean up, and capillary gas chromatographic analysis J. High Resolution Chromatography, 13, 84-98 (1990) Patent documents US 4726954; US4636468; US2005059130; EP2290059; EP3081644; EP2254996; EP1776455

[0169] Sequence Listing >SEQ ID NO:1 MVLSTVIGEWFSRVLFGTVAPSPLTATAPISQDFYDTALTFSHLSNVAYCINTPLESLKSDFSCGVACSHFPNMELVEEFGGEFFETSITGFLSIDHVKKEKYVVYRGTYDIGDVYTDIQLSQSPFLVTPSALGSTANLCEGCTIHDGWNKAYNETMGIIGDKLADHVNSNPDY RLVVTGHSLGAAIAVLSATSLKVNGQDPYLYTYGQPRIGNANFANFVSKQWFGEGDGLSMDSDRRYFRLTHWNDLFVGFPAFKDYVHSVGEIYIDYFTVQPPLNKVFSCAGPESMSCYRKDFNALARLDIVKNHLAYFDWISLCTLNIGRRDLERGRKFEGTWLYGGLANGSTIF >Sequence number 2 AVLQKRVYTSTETSHIDQESYNFFEKYARLANIGYCVGPGTKIFKPFNCGLQCAHFPNVELIEEFHDPRLIFDVSGYLAVDHASKQIYLVIRGTHSLEDVITDIRIMQAPLTNFDLAANISSTATCDDCLVHNGFIQSYNNTYNQIGPKLDSV IEQYPDYQIAVTGHSLGGAAALLFGINLKVNDHDPLVVTLGQPIVGNAGFANWVDKLFFGQENPDVSKVSKDRKLYRITHRGDIVPQVPFWDGYQHCSGEVFIDWPLIHPPLSNVVMCQGQSNKQCSAGNTLLQQVNVIGNHLQYFVTEGVCGI > Sequence number 3 QTAPITQETYDFVLKYGWLSNVAYCVRAPGPFALQSDFTCGNSCAHFPDVTLDYQFGGNFFSTSVTGFLAHDHTKKEKYIVFRGTFSIADAITDIQTIQQPYMTSIPPLNTTDINSTNPSASINCPGCQVHDGFQKAYRETMVNVQDRLVDFLGNNTDYKLIVTGHSLGAVTA LFMGINLKNLGYDPTLINYGQPRLGNKAFADYVDALFFKQGDDGLTINPERRMYRVTHWNDFFVGWPAGYSHTLGEVYISDPTGINAPIEDVYSCAGPENNQCHHGSFNLLERLNILKNHCGYLNWIFYCAINVDKREMMIDPPRVGKRVEHWSGKFSDVESTEGLMYEAIYPM > sequence number 4 APAPAPMQRRDISSTVLDNIDLFAQYSAAAYCSSNIESTGTTLTCDVGNCPLVEAAGATTIDEFDDSSSYGDPTGFIAVDPTNELIVLSFRGSSDLSNWIADLDFGLTSVSSICDGCEMHKGFYEAWEVIADTITSKVE AAVSSYPDYTLVFTGHSYGAALAAVAATVLRNAGYTLDLYNFGQPRIGNLALADYITDQNMGSNYRVTHTDDIVPKLPPELLGYHHFSPEYWITSGNDVTVTTSDVTEVVGVDSTAGNDGTLLDSTAHRWYTIYISECS > sequence number 5 VPMLQSRATSDPAEWTELHRAAQLSSAAYTGCTGSAFDVTITKQINELVTDTQGFIGYSTEKKRITVAMRGSTSATDIANDVDTTLVEPTLSGVNFPSGAKMMHGIYSPWSSVHDDVISEVKSLVEQYPDYTIESTGHSLGGSLTYISYIALAQNFPGKTIISNALAAYPIGNEAFANFGASQNGTLNRGNNADDGVPNMYVMWPWDFVHYGTEYYSSGTQASTVKCSGERDTSCSAGNGQVGVTAGHFSNFGIAMGMAGC > Sequence number 6 APASNDATIADVSSTFTLPPIIKDRVAFSNDIADTDYFNLTKRASETTVGGNTMDLPSNAPALPAVPKAGDVVIATAAQIAEYKKYAALASTAYCRSVVPLNLWTCVNCLRFAPDGKLIKTFSSVISDTNGFVLRSDAQKTIYVVFRGTNSIRSAITDLVFTLISYP PVSGARVHTGFYASYQAVVSDYFPVVQSQLTSYPSYKVVVTGHSLGGAHALLAGMDLYQREKRLTASNLFIHTAGCPRVGNPTFANYVASTGITFTRSVNQRDIVPHVPPTYAGYLHPGVEVWARTSSTVQICTSNTESNMCSNSIEPFTSFTDHLSYYGITEGVCI > Sequence number 7 SPVQLARRAISSELLERFTLFSQFATLSACDQNINHTGQSLTCDYGTCGLVAADNTTVINAFHSDNGPTGYIALDHTRQLIVLTFRGTVSKSDGDTDLDIVLTSIDDVCTGCKAHHGFWVYWSAVASQATTQLQDATSAYPSY RLSVVGHSLGGGIAALAGTVLRTQGFTLDIWTFGGPKPGNLKLAEFITNQQPPNSIYRATHTTDPIPKVPLNLPFLDWSQPSPEYWITQETGVQVTTDGVEYIEGINSKAGNAGSDRDLRWPNPEHGWYFGNMSVCASPSDASS > Sequence number 8 APSQLVPRAVSSGTLDQLTLFAQYSAASYCANNVNSPGDAITCSGGYCPKLQSAGVKSLFEFDDSTEFGDVAGFLSVDTANKLLVLSFRGSRTISNWIANLDFGQADASSLCIGCKAHSGFLKAWTVVSDDVMPPLV SAMAKYPGFRLVLTGHSFGGAVAALGATALRKAGYKLDLYTYGQPRVGNTALATYMTNQGSMYRVTHSNDIVPKLPPPLLGYTHASPEYWITSGDNVAVTTKDITQVNGIGSKDGNAGSNGDSIPAHNWYIVNIDGCK > Sequence number 9 APQKRSVSSTVLAQLSRYAQWSAAAYCSGNTSGANQVVSCSANNCPDVQASGATMLYEFDSTNTYGDAAGFLAADTTQQQIILSFRGSRTTSNWIANLDTELTSSTLCSGCEVHQGFWLDYQTVAATLKAQIDAA LNTYPGYSLVVTGHSLGGALAMLAGLDLNSQGYAPTIYTYGQPRVGNLALAQYITNVGNQWRVTHADDAVPKLPPRLFGFSHASPEYWITSGDNVAVTTGDVTVVTGVNSLGGNDGTLTASVSAHNWYLVDIDACK > Sequence number 10 APALPLEVRDSGVSQAVYDDLVVYAKYSSAVYQPFCPRPLGSWMIKAFDTKGTQGFVARDDARKEIIVAFRGSFEIVDILIDIQIILTPLSTPGVSNVGSARVHTGFQKAYNFVIDEVQSLMKSQIDAYPSYKL VVTGHSLGGAVATFAALALKSKYPSKSLRLFTYGAPRVGDAAFASLVESRLGINNIYRGVHTWDGVPTLLGRWLGYRHYGTEYWQHKDPSKPENVRKCNGGEDTSCSNSIISTGINPPHAFQFGQVMAINPLLCF >SEQ ID NO: 11 MVSFGARIKDFFSVLLFGAASTSSSTKTALVSQGFYDAALDFSHLSNIAYCVNAPITPLKSDFSCGQSCVHFPDIELVHIFGGDFFSTSITGYLALDHVKKEKYVVFRGTFSIADAITDIQFQQSSFLVNVPALNTFTANDTAPEAQIDCKQCKIHDGFSKAFTETWHNIGDLLEQHLDSYPD YQLYVTGHSLGAAMALLGATSIKLRGYDPILINYGQPRVGNKAFADYISALWFGNGDGLEINQQRRLYRMTHWNDVFVGLPNWDGYTHSNGEVYIKGKYVNPPLKDVFSCAGGENSKCYRSEFNLLAQINLLQNHLCYIDYIGFCALNVGRRELNDLPHYNGPYKYGHKTEEQFIAEGLELSN >SEQ ID NO: 12 >> GCTGTTTTGCAAAAGAGAGTTTACACTTCTACTGAAACTTCTCATATCGATCAAGAATCTTACAACTTTTTCGAGAAGTATGCTAGATTGGCTAATATTGGTTATTGTGTTGGTCCAGGTACTAAGATTTTTAAGCCTTTCAACTGTGGTTTGCAATGTGCTCATTTCCCAAACGTTGAATTGATCGAAGAGTTTCACGATCCTAGATTGATTTTCGATGTTTCTGGTT CTTGGCTGTTGATCATGCTTCTAAGCAAATCTACTTGGTTATTAGAGGTACTCACTCTTTGGAGGATGTTATCACTGATATCAGAATCATGCAAGCTCCATTGACTAACTTTGATTTGGCTGCTAATATTTCTTCTACTGCTACTTGTGATGATTGTTTGGTTCACAACGTTTCATTCAATCTTACAACAACACTTACAATCAAATTGGTCCAAAGTTGGATTCTGTTA TTGAGCAATACCCTGATTATCAAATTGCTGTTACTGGTCATTCTTTGGGTGGTGCTGCTGCTTTGTTGTTCGGTATTAACTTGAAGGTTAATGATCACGATCCATTGGTTGTTACTTTGGGTCAACCTATTGTTGGTAACGCTGGTTTCGCTAATTGGGTTGATAAGTTGTTTTTCGGTCAAGAAAACCAGATGTTTCTAAGGTTTCTAAGGATAGAAAGTTGTTACAGA ATCACTCATAGAGGAGATATTGTTCCACAAGTTCCTTTTTGGGATGGTTATCAACATTGTTCTGGAGAGGTTTTCATTGATTGGCCTTTGATTCACCCACCTTTGTCTAATGTTGTTATGTGTCAAGGTCAATCTAACAAACAATGTTCTGCTGGTAAACACTTTGTTGCAACAAGTTAACGTTATTGGTAATCACTTGCAATACTTCGTTACTGAAGGTGTTTGTGGTATT > அக்கை >> GCTCCAGCTCCTGCTCCAATGCAAAGAAGAGATATCTCTTCTACTGTTTTGGATAACATCGATTTGTTCGCTCAATACTCTGCTGCTGCTTATTGTTCTCTAACATTGAATCTACTGGTACTACTTTGACTTGTGATGTTGGTAATTGTCCTTTGGTTGAAGCTGCTGGTGCTACTACTATCGATGAGTTCGATGATTCTTCTTCTTA CGGAGATCCTACTGGTTTCATTGCTGTTGATCCAACTAACGAGTTGATTGTTTGTCTTTTAGAGGTTCTTCTGATTTGTCTAATTGGATTGCTGATTTGGATTTCGGTTTGACTTCTGTTTCTTCTATTTGTGATGGTTGTGAAATGCATAAGGGTTTTACGAAGCTTGGGAGGTTATTGCTGATACTATCACTTCTAAGGTTGAAG CTGCTGTTTCTTCTTACCCTGATTATACTTGGTTTTCACTGGTCACTCTTATGGTGCTGCTTTGGCTGCTGTTGCTGCTACTGTTTTGAGAAATGCTGGTTACACTTTGGATTTGTATAACTTTGGTCAACCAAGAATTGGTAATTTGGCTTTGGCTGATTACATCACTGATCAAAACATGGGTTCTAACTACAGAGTTACTCATACT GATGATATTGTTCCTAAGTTGCCACCTGGAATTGTTGGGTTACCATCACTTTTCTCCAGAGTATTGGATTACTTCTGGTAACGATGTTACTGTTACTACTTCTGATGTTACTGAAGTTGTTGGTGTTGATTCTACTGCTGGTAATGATGGTACTTTGTTGGATTCTACTACTGCTCACAGATGGTACACTATCTACATTTCTGAGTGTTCT > அக்கை GTTCCAATGTTGCAATCTAGAGCTACTTCTGATCCTGCTGAATGGACTGAGTTGCATAGAGCTGCTCAATTGTCTTCTGCTGCTTACACTGGTTGTACTGGTTCTGCTTTCGATGTTACTATTCACTAAGCAAATTAACGAATTGGTTACTGATACTCAAGGTTTCATTGGTTACTCTACTGAGAAGAAAAGAATC ACTGTTGCTATGAGAGGTTCACTTCTGCTACTGATATTGCTAACGATGTTGATACTACTTTGGTTGAACCAACTTTGTCTGGTGTTAATTTTCCTTCTGGTGCTAAGATGATGCATGGTATCTACTTCCATGGTCTTCTGTTCACGATGATTTATCTCTGAGGTTAAGTCTTTGGTTGAGCAATACCCTGATT ATACTATTGAATCTACTGGTCACTCTTTGGGTGGTTCTTTGACTTACATCTCTTACATCGCTTTGGCTCAAAAACTTCCCAGGTAAAACTATCATCTCTAACGCTTTGGCTGCTTATCCTATTGGTAACGAGGCTTTTGCTAATTTCGGTGCTTCTCAAAACGGTACTTTGATCGTGGTAACAATGCTGATGATGG TGTTCCAAATATGTACGTTATGTGGCCTTGGGATTTCGTTCATTACGGTACTGAATACTACTCTTCTGGTACTCAAGCTTCTACTGTTAAATGTTCTGGAGAGAGATACTTCTTGTTCTGCTGGTAACGGTCAAGTTGGTGTTACTGCTGGTCACTTTTCTAATTTCGGTATTGCTATGGGTATGGCTGGTTGTT >> GCTCCTGCTTCTAATGATGCTACTATTGCTGATGTTTCTTCTACTTTCACTTTGCCACCTATTATTAAGGATAGAGTTGCTTTTTCTAACGATATCGCTGATACTGATTACTTCAACTTGACTA AGAGAGCTTCTGAAACTGTTGGTGGTAATACTATGGATTTGCCATCTAACGCTCCAGCTTTGCCTGCTGTTCCAAAGGCTGGAGATGTTGTTATTGCTACTGCTGCTCAAATTGCTGAGTACAAG AAATATGCTGCTTTGGCTTCTACTGCTTACTGTAGATCTGTTGTTCCTTTGAATTTGTGGACTTGGTGTTAACTGTTTGAGATTTGCTCCAGATGGTAAATTGATTAAAACTTTCTCTTCTGTTAT TTCTGATACTAACGGTTTCGTTTTGAGATCTGATGCTCAAGACTATCTACGTTGTTTTCAGAGGTACTAACTCTATCAGATCTGCTATCACTGATTTGGTTTTTACTTTGATCTCTTACCCAC CTGTTTCTGGTGCTAGAGTTCATACTGGTTTCTACGCTTCTTATCAAGCTGTTGTTTCTGATTACTTTCCTGTTGTTCAATCTCAATTGACTTCTTACCCATCTTATAAGGTTGTTGTTACTGGTCATTCTTTGGGTGGTGCTCACGCTTTGTTGGCTGGTATGGATTTGTACCAAAGAGAAAAGAGATTGACTGCTTCTAATTTGTTTATTCACACTGCTGGTTGTCCTAGAGTTGGTAATCCAACTTTCGCTAACTACGTTGCTTCTACTGGTATCACTTTTACTAGATCTGTTAACCAAAGAGATATTGTTCCTCATGTTCCACCTACTTACGCTGGTTATTTGCACCCAGGTGTTGAAGTTTGGGCTAGAACTTCTTCTACTGTTCAAATCTGTACTTCTAACACTGAATCTAACATGTGTTCTAACTCTATCGAGCCTTTTACTTCTTTCACTGATCATTTGTCTTACTATGGTATTACTGAGGGTGTTTGTATT > அக்கை TCTCCAGTTCAATTGGCTAGAAAGAGCTATTTCTTCTGAATTGTTGGAGAGATTCACTTTGTTCTCTCAATTGCTACTTTGTCTGCTTGTGAATCAAAACATCAACCATACTGGTCAATCTTTGACTTGTGATTACGGTACTTGTGGTTTGGTTGCTGCTGATAACACTACTGTTATTAATGCTTTCCATTCTGATAACGGTCCTACTGGTTATAT TGCTTTGGATCACACTAGACAATTGATCGTTTTGACTTTTAGAGGTACTGTTTCTAAGTCTGATGGAGATACTGATTTGGATATCGTTTTGACTTCTATCGATGATGTTTGTACTGGTTGTAAAGCTCATCACGGTTTCTGGGTTTACTGGTCTGCTGTTGCTTCTCAAGCTACTCAATTGCAAGATGCTACTTCTGCTTACCCATCTTATA GATTGTCTGTTGTTGGTCATTCTTTGGGTGGTGGTATTGCTGCTTTGGCTGGTACTGTTTTGGAGAACTCAAGGTTTCACTTTGGATATTTGACTTTTGGTGGTCCAAAGCCTGGTACTTGAAGTTGGCTGAATTCATTACTAACCAACAACCACCTAATTCTATCTACAGAGCTACTCACACTACTGATCCAATTCCTAAGGTTCCATTGAAT TTGCCATTTTTGGATTGGTCTCAACCATCTCCTGAATACTGGATTACTCAAGAGACTGGTGTTCAAGTTACTACTGATGGTGTTGAATACATCGAGGGTATTAACTCTAAAGCTGGTAATGCTGGTTCTGATAGAGATTTGAGATGGCCAAACCCTGAGCACGGTTGGTATTTTGGTAATATGTCTGTTTGTGCTTCTCCTCTGATGCTTCTTCT > அக்கை GCTCCTTCTCAATTGGTTCCAAGAGCTGTTTCTTCTGGTACTTTGGATCAATTGACTTTGTTTGCTCAATACTCTGCTGCTTCTTATTGTGCTAACAATGTTAACTCTCCTGGAGATGCTATTACTTGTTCTGGTGGTTACTGTCCAAAGTTGCAATCTGCTGGTGTTAAGTCTTTGTTCGAATTCGATGATTCTACTGAGTTTGG AGATGTTGCTGGTTTCTTGTCTGTTGATACTGCTAATAAGTTGTTGGTTTTGTCTTTTAGAGGTTCTAGAACTATCTCTAACTGGATCGCTAATTTGGATTTCGGTCAAGCTGATGCTTCTTCTTTGTGTATTGGTTGTAAGGCTCATTCTGGTTTCTTGAAAGCTTGGACTGTTGTTTCTGATGATGTTATGCCACCTTTGGTTT CTGCTATGGCTAAATACCCTGGTTTTTAGATTGGTTTTGACTGGTCACTCTTTCGGTGGTGCTGTTGCTGCTTTGGGTGCTACTGCTTTGAGAAAGGCTGGTTACAAGTTGGATTTGTACACTTACGGTCAACCAAGAGTTGGTAACACTGCTTTGGCTACTTACATGACTAATCAAGGTTCTATGTACAGAGTTACTCATTCTAAC GATATCGTTCCTAAGTTGCCACCTCCATTGTTGGGTTACACTCACGCTTCTCCAGAATATTGGATTACTTCTGGAGATAACGTTGCTGTTACTACTAAGGATATCACTCAAGTTAACGGTATCGGTTCTAAAGATGGTAACGCTGGTTCTAATGGAGATTCTATTCCTGCTCATAACTGGTATATTGTTAATATTGATGGTTGTAAA > அக்கை GCTCCACAAAAGAGATCGTTTCTTCTACTGTTTTGGCTCAATTGTCTAGATACGCTCAATGGTCTGCTGCTGCTTATTGTTCTGTAACACTTCTGGTGCTAATCAAGTTGTTTCTTGTTCTGCTAACAATTGTCCTGATGTTCAAGCTTCTGGTGCTACTATGTTGTACGAATTCGATTCTACTAACACTTACGGAGATGC TGCTGGTTTCTTGGCTGCTGATACTACTCAACAAAATCATCTTGTCTTTTAGAGGTTCTAGAACTACTTCTAACTGGATTGCTAATTTGGATACTGAATTGACTTCTTCTACTTTGTGTTCTGGTTGTGAGGTTCATCAAGGTTTCTGGTTGGATTACCAAACTGTTGCTGCTACTTTGAAAGCTCAAATTGATGCTGCTT TGAATACTTACCCAGGTTATTCTTTGGTTGTTACTGGTCACTCTTTGGGTGGTGCTTTGGCTATGTTGGCTGGTTTGGATTTGAACTCTCAAGGTTACGCTCCAACTATCTACACTTATGGTCAACCTAGAGTTGGTAATTTGGCTTTGGCTCAATACATCACTAACGTTGGTAACCAATGGAGAGTTACTCATGCTGATGAT GCTGTTCCAAAGTTGCCACCTAGATTGTTTGGTTTCTCTCACGCTTCTCCTGAGTACTGGATTACTTCTGGAGATAACGTTGCTGTTACTACTGGGATGTTACTGTTGTTACTGGTGTTAACTCTTTGGGTGGTAATGATGGTACTTTGACTGCTTCTGTTTCTGCTCATAACTGGTATTTGGTTGATATCGATGCTTGTAAA > அக்கை GCTCCAGCTTTGCCTTTGGAAGTTAGAGATTCTGGTGTTTCTCAAGCTGTTTACGATGATTTGGTTGTTTACGCTAAGTATTCTTCTGCTGTTTATCAACCATTTGTCCAAGAACCTTTGGGTTCTTGGATGATTAAGGCTTTTGATACTAAAGGTACTCAAGGTTTCGTTGCTAGAGATGATGCTAGAAAGGAAATCATC GTTGCTTTTAGAGGTTCTTTCGAGATTGTTGATATCTTGATCGATATCCAAATCATCTTGACTCCATTGTCTACTCCTGGTGTTTCTAACGTTGGTTCTGCTAGAGTTCATACTGGTTTCCAAAAGGCTTACAACTTCGTTATCGATGAGGGTTCAATCTTTGATGAAGTCTCAAATCGATGCTTACCCTTCTTACAAGTTGG TTGTTACTGGTCACTCTTTGGGTGGTGCTGTTGCTACTTTCGCTGCTTTGGCTTTGAAGTCTAAGTACCCATCTAAGTCTTTGAGATTGTTTACTTACGGTGCTCTAGAGTTGGAGATGCTGCTTTCGCTTCTTGGTTGAATCTAGATTGGGTATTAACAACATCTACAGAGGTGTTCACTTGGGATGGTGTTCCAAC TTTGTTGGGTAGATGGTTGGGTTACAGACATTATGGTACTGAGTATTGGCAACACAAGGATCCATCTAAACCTGAAAACGTTAGAAAGTGTAATGGTGGAGAGGATACTTCTTGTTCTAACTCTATCATCTCTACTGGTATTAATCCACCTCACGCTTTTCAATTCGGTCAAGTTATGGCTATTAACCCTTTGTTGTGTTTT > அக்கை >SEQ ID NO: 23

Claims

1. 1. A method for preparing a food product, comprising the steps of: using a lipase comprising an amino acid sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to SEQ ID NO:1 or SEQ ID NO:3; or using a DNA sequence encoding a lipase selected from sequences having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 12 or SEQ ID NO:

14. Including, where: The method, wherein the food product is a dairy product selected from cheese, processed cheese, cheese-like products, enzyme-treated cheese, butter, yogurt, cream, or seasonings.

2. 2. The method of claim 1, wherein the DNA sequence is an isolated DNA sequence, a recombinant DNA sequence, or a synthetic DNA sequence.

3. The method of claim 1 or 2, wherein the DNA sequence comprises a signal peptide sequence.

4. The lipase is C 10 -When compared to the release of fatty acids, C 8 -When compared to the release of fatty acids, C 6 -When compared to the release of fatty acids, C 12 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 18:0 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 16 -relative to the release of fatty acids, or C 8 -Fatty acid~C 18:2 -When compared to the release of fatty acids, C 10 -Fatty acid~C 18:1 -When compared to the release of fatty acids, C 12 -Fatty acid~C 18:0 -When compared to the release of fatty acids, C 14 -Fatty acid~C 16:0 -relative to the release of fatty acids, or C 16 ~C 18 -C from dairy compositions containing milk fat and / or other fats when compared to the release of fatty acids 4 - A method according to any one of claims 1 to 3, which has a higher specificity for the release of fatty acids.

5. The lipase is C 10 -compared to the release of fatty acids, or C 6 -When compared to the release of fatty acids, C 8 -When compared to the release of fatty acids, C 12 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 16 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 18:3 -relative to the release of fatty acids, or C 16 ~C 18 -C from dairy compositions containing milk fat and / or other fats when compared to the release of fatty acids 4 - A method according to any one of claims 1 to 4, which has a higher specificity for the release of fatty acids.

6. The lipase is C at pH 6.6 to 6.8 or below pH 6. 10 -C from dairy compositions containing milk fat and / or other fats when compared to the release of fatty acids 4 - The method according to claim 5, which has a higher specificity for the release of fatty acids.

7. The lipase is C at pH 3.8 to 5.

6. 10 -C from dairy compositions containing milk fat and / or other fats when compared to the release of fatty acids 4 - The method according to claim 6, which has a higher specificity for the release of fatty acids.

8. The lipase is C at pH 4.4 to 5.

4. 10 -C from dairy compositions containing milk fat and / or other fats when compared to the release of fatty acids 4 - The method according to claim 7, which has a higher specificity for the release of fatty acids.

9. The lipase is C at pH 4.6 to 5.

2. 10 -C from dairy compositions containing milk fat and / or other fats when compared to the release of fatty acids 4 - The method according to claim 8, which has a higher specificity for the release of fatty acids.

10. The lipase is C at a temperature of less than 20°C. 10 -compared to the release of fatty acids, or C 6 -When compared to the release of fatty acids, C 8 -When compared to the release of fatty acids, C 12 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 16 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 18:3 -relative to the release of fatty acids, or C 16 ~C 18 -C from dairy compositions containing milk fat and / or other fats when compared to the release of fatty acids 4 - A method according to any one of claims 1 to 9, which has a higher specificity for the release of fatty acids.

11. The lipase is 10 -C from dairy compositions containing milk fat and / or other fats when compared to the release of fatty acids 4 - A method according to any one of claims 1 to 10, which has a higher specificity for the release of fatty acids.

12. The lipase is C at a temperature of less than 10°C. 10 -C from dairy compositions containing milk fat and / or other fats when compared to the release of fatty acids 4 - A method according to any one of claims 1 to 11, which has a higher specificity for the release of fatty acids.

13. The lipase is C at a temperature of 5 to 8°C. 10 -C from dairy compositions containing milk fat and / or other fats when compared to the release of fatty acids 4 - A method according to any one of claims 1 to 12, which has a higher specificity for the release of fatty acids.

14. The method according to any one of claims 1 to 13, wherein the lipase is a microbial lipase.

15. 15. The method of claim 14, wherein the microbial lipase is an isolated microbial lipase, a recombinant microbial lipase, or a synthetic microbial lipase.

16. 16. The method according to any one of claims 1 to 15, wherein the dairy product is a cheese selected from Feta cheese, Provolone cheese, Pecorino cheese, Parmesan cheese, Grana Padano cheese, Parmigiano-Reggiano cheese, Romano cheese, Chester cheese, Dumbo cheese, Manchego cheese, Saint Paulin cheese, Cheddar cheese, Monterey cheese, Colby cheese, Edam cheese, Gouda cheese, Muenster cheese, Swiss-type cheese, Gruyere cheese, or Emmental cheese; or Pasta Filata cheese; or Fresh cheese; or Cream cheese; or White mould cheese; or Blue mould cheese.

17. The method of claim 16, wherein the pasta filata cheese is mozzarella or queso fresco; the fresh cheese is ricotta, cream cheese, Neuchâtel or cottage cheese; the white mould cheese is Brie or Camembert; and the blue mould cheese is Gorgonzola or Danish blue cheese.

18. using a lipase comprising an amino acid sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 1 or SEQ ID NO: 3, or using a DNA sequence encoding a lipase selected from sequences having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to SEQ ID NO: 12 or SEQ ID NO: 14, 1. A method of producing a food product, comprising: The method, wherein the food product is a dairy alternative.

19. 20. The method of producing a food product according to claim 18, wherein the DNA sequence is an isolated DNA sequence, a recombinant DNA sequence or a synthetic DNA sequence.

20. 19. The method of claim 18, wherein the dairy alternative product is a dairy alternative cheese or a vegan cheese.

21. The lipase is C 10 -When compared to the release of fatty acids, C 8 -When compared to the release of fatty acids, C 6 -When compared to the release of fatty acids, C 12 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 18:0 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 16 -When compared to the release of fatty acids, C 8 -Fatty acid~C 18:2 -When compared to the release of fatty acids, C 10 -Fatty acid~C 18:1 -When compared to the release of fatty acids, C 12 -Fatty acid~C 18:0 -relative to the release of fatty acids, or C 14 -Fatty acid~C 16:0 -When compared to the release of fatty acids, C 16 -C 18 C from lipid-containing compositions or from water-in-oil emulsions when compared to the release of fatty acids 4 - A method according to any one of claims 18 to 20, which has a higher specificity for the release of fatty acids.

22. The method described in claim 21, wherein the lipid-containing composition is a vegetable fat-containing composition.

23. A food product comprising a lipase, wherein the lipase comprises an amino acid sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to SEQ ID NO:1 or SEQ ID NO:3; and wherein the food product is C 6 -Fatty acids, C 8 -Fatty acids, C 10 -Fatty acids, C 12 -Fatty acids, C 14 -Fatty acids, C 16 -Fatty acids, C 18:1 -Fatty acids, C 18:2 -fatty acid or C 18:3 -More C than other fatty acids selected from fatty acids 4 -contains fatty acids, and / or wherein the food product is C 8 -Fatty acids, C 10 -Fatty acids, C 12 -Fatty acids, C 14 -Fatty acids, C 16 -Fatty acids, C 18:1 -Fatty acids, C 18:2 -fatty acid or C 18:3 -More C than other fatty acids selected from fatty acids 6 -contains fatty acids, The food product is a dairy product selected from cheese, processed cheese, enzyme-treated cheese, cheese-like products, butter, yogurt, cream, seasonings, or a dairy alternative product.

24. The food product of claim 23, wherein the dairy alternative product is a dairy alternative cheese or a vegan cheese.

25. The cheese is feta cheese, provolone cheese, pecorino cheese, parmesan cheese, grana padano cheese, parmigiano reggiano cheese, romano cheese, chester cheese, dambo cheese, manchego cheese, saint paulin cheese, cheddar cheese, monterey cheese, colby cheese, edam cheese, gouda cheese, muenster cheese, swiss type cheese, gruyere cheese or emmental cheese; or pasta filata cheese; or fresh cheese; or cream cheese; or white mold cheese; or blue mold cheese; 24. A food product comprising the lipase of claim 23.

26. 26. The food product comprising the lipase of claim 25, wherein the cheese is feta cheese or provolone cheese.

27. ​​The food product of claim 25, wherein the pasta filata cheese is mozzarella or queso fresco; the fresh cheese is ricotta, cream cheese, Neuchâtel or cottage cheese; the white mould cheese is brie or camembert; and the blue mould cheese is gorgonzola or Danish blue cheese.

28. 1. Use of a lipase for producing a food product, comprising: wherein the food product is a dairy product selected from cheese, processed cheese, enzyme-treated cheese, cheese-like products, butter, yogurt, cream, or seasonings; or wherein the food product is a dairy alternative; and Here, the use of a lipase, wherein the lipase comprises an amino acid sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to SEQ ID NO:1, SEQ ID NO:

3.

29. The use described in claim 28, wherein the dairy alternative product is a dairy alternative cheese or a vegan cheese.

30. The lipase is C 10 -When compared to the release of fatty acids, C 8 -When compared to the release of fatty acids, C 6 -When compared to the release of fatty acids, C 12 -When compared to the release of fatty acids, C 18:2 -When compared to the release of fatty acids, C 14 -When compared to the release of fatty acids, C 18:0 -When compared to the release of fatty acids, C 18:1 -When compared to the release of fatty acids, C 16 -relative to the release of fatty acids, or C 8 -Fatty acid~C 18:2 -When compared to the release of fatty acids, C 10 -Fatty acid~C 18:1 -When compared to the release of fatty acids, C 12 -Fatty acid~C 18:0 -When compared to the release of fatty acids, C 14 -Fatty acid~C 16:0 -relative to the release of fatty acids, or C 16 ~C 18 -C from dairy compositions containing milk fat and / or other fats when compared to the release of fatty acids 4 - Use of a lipase according to claim 28, which has a higher specificity for the release of fatty acids.

31. A milk composition comprising milk fat and / or other lipids, C 8 -Fatty acids, C 10 -Fatty acids, C 12 -Fatty acids, C 14 -Fatty acids, C 16 -Fatty acids, C 18:0 -Fatty acids, C 18:1 -Fatty acids, C 18:2 -fatty acids, or C 18:3 -C due to the activity of producing or releasing fatty acids 4 -fatty acids and / or C 6 - Use of a lipase with a higher activity of producing or releasing fatty acids, wherein the lipase comprises an amino acid sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to SEQ ID NO:1 or SEQ ID NO:

3.

32. In food products, C 8 -Fatty acids, C 10 -Fatty acids, C 12 -Fatty acids, C 14 -Fatty acids, C 16 -Fatty acids, C 18:0 -Fatty acids, C 18:1 -Fatty acids, C 18:2 -fatty acids, or C 18:3 -C in the food product due to the activity of producing or releasing fatty acids 4 -fatty acids and / or C 6 - the use of lipases with a higher activity of producing or releasing fatty acids, wherein said lipase comprises an amino acid sequence having at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% identity with SEQ ID NO: 1 or SEQ ID NO: 3, Use of lipase.

33. the food product is a dairy product selected from cheese, processed cheese, enzyme-treated cheese, cheese-like products, butter, yogurt, cream, or seasonings; or 33. The use according to claim 32, wherein the food product is a dairy alternative product.

34. The use described in claim 33, wherein the dairy alternative product is a dairy alternative cheese or a vegan cheese.

35. 34. The use according to claim 32 or 33, wherein the food product is a dairy product selected from cheese, and wherein the cheese is Feta cheese, Provolone cheese, Pecorino cheese, Parmesan cheese, Grana Padano cheese, Parmigiano-Reggiano cheese, Romano cheese, Chester cheese, Dumbo cheese, Manchego cheese, Saint Paulin cheese, Cheddar cheese, Monterey cheese, Colby cheese, Edam cheese, Gouda cheese, Muenster cheese, Swiss-type cheese, Gruyere cheese, or Emmental cheese; or Pasta Filata cheese; or Fresh cheese; or Cream cheese; or White mould cheese; or Blue mould cheese.

36. 36. The use of claim 35, wherein the cheese is feta or provolone cheese.

37. The use described in claim 35, wherein the pasta filata cheese is mozzarella or queso fresco cheese; the fresh cheese is ricotta, cream cheese, Neuchâtel or cottage cheese; the white mould cheese is brie or camembert cheese; and the blue mould cheese is gorgonzola or Danish blue cheese.

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