Glutaminase and methods of use thereof

Abstract

The present invention relates to glutaminases and their use in the manufacture of flavors for incorporation in foods. More particularly, the present invention relates to methods of making glutamate from protein sources and the use of such glutamate in foods.

Description

[0001] NB41870-WO-PCT[3] TITLE GLUTAMINASE AND METHODS OF USE THEREOF TECHNICAL FIELD The present invention relates to glutaminases and their use in the manufacture of flavors for incorporation in foods. More particularly, the present invention relates to methods of making glutamate from protein sources and the use of such glutamate in foods. REFERENCE TO A SEQUENCE LISTING The contents of the electronic submission of the text file Sequence Listing, named “NB41870WOPCT3_SequenceListing” was created on September 12, 2024, and is 40 KB in size, which is hereby incorporated by reference in its entirety. BACKGROUND Mono sodium glutamate (MSG) is a popular seasoning and provides a strong umami or savoury flavour when incorporated into foods. MSG typically comes as a white, crystalline powder. MSG is commonly added to food to provide a savoury flavour. Examples of such food include bouillon cubes, soups, stews, gravy, condiments and various savoury snacks. While MSG is generally considered safe to eat, some consumers claim exogenously added MSG causes headaches. In this regard, many consumers have a negative perception of foods to which exogenous MSG has been added. Further, some jurisdictions require labelling of food products to which exogenous MSG has been added. As of today, most of the world’s MSG is made by fermentation. A strain of Corynebacterium glutamicum is used to convert glucose and ammonia into glutamic acid. Glutamic acid is neutralized via sodium hydroxide to produce MSG which is then extracted from the fermentate and purified to provide the commercial MSG product. There is a continuing need in the art for MSG which is derived from natural ingredients and does not require labelling. NB41870-WO-PCT[3] SUMMARY OF THE INVENTION In an aspect of the present invention, a method for producing a hydrolysate enriched in glutamate is presented having the step of adding a glutaminase corresponding to an isolated polypeptide having at least 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof to a starting material comprising glutamine and incubating for a sufficient time to produce the hydrolysate. Optionally, the isolated polypeptide has at least 80% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has at least 85% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has at least 90% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has at least 95% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has at least 98% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has at least 99% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the step of incubating is conducted at a temperature of around 40 to 70°C. Optionally, the step of incubating is conducted at a temperature of around 45 to 65°C. Optionally, the step of incubating is conducted at a temperature around 50 to 60°C. Optionally, the step of incubating is conducted at a temperature around 55°C. NB41870-WO-PCT[3] Optionally, the starting material is a plant protein. Optionally, the plant protein is selected from the group consisting of pea, soy, fava, gluten, and oat. Optionally, the method has the further step of adding an exo-peptidase. Optionally, the exo-peptidase is an isolated polypeptide having at least 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:23 or an exopeptidase active fragment thereof. Optionally, the method has the further step of adding an endo-peptidase. Optionally, the endopeptidase is an isolated polypeptide having at least 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:22 or an endopeptidase active fragment thereof or SEQ ID NO:24 or an endopeptidase active fragment thereof. In another aspect of the present invention, a protein hydrolysate enriched in glutamate is presented which is produced according to the method described above. In another aspect of the present invention, use of a protein as made above to produce a broth, a meat product or a cheese is presented. In another aspect of the present invention, a method of providing umami flavor to a plant-based meat alternative is presented having the steps of: a. providing a batter comprising a plant protein; b. combining the batter with a glutaminase enzyme comprising an isolated polypeptide having at least 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof; c. incubating the batter at a temperature of about 40 to 70°C; and d. cooking the batter to provide the plant-based meat alternative. Optionally, the isolated polypeptide has at least 80% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has at least 85% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has at least 90% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has NB41870-WO-PCT[3] at least 95% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has at least 98% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has at least 99% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the step of incubating is conducted at a temperature of around 40 to 70°C. Optionally, the step of incubating is conducted at a temperature of around 45 to 65°C. Optionally, the step of incubating is conducted at a temperature around 50 to 60°C. Optionally, the step of incubating is conducted at a temperature around 55°C. Optionally, the plant protein is selected from the group consisting of pea, soy, fava, gluten, and oat. Optionally, has the further step of adding an exo-peptidase at step a and / or step b. Optionally, the exo-peptidase is an isolated polypeptide having 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:23 or a proteolytically active fragment thereof. Optionally, the method has the further step of adding an endo-peptidase at step a and / or step b. Preferably, the endo-peptidase comprises an isolated polypeptide having 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:22 or SEQ ID NO:24 or an exopeptidase active fragment thereof. In another aspect of the present invention, a recombinant glutaminase is presented which is an isolated polypeptide having at least 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof to a starting material comprising glutamine and incubating for a sufficient time to produce the hydrolysate. Optionally, the isolated polypeptide has at NB41870-WO-PCT[3] least 80% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has at least 85% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has at least 90% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has at least 95% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has at least 98% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has at least 99% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Optionally, the isolated polypeptide has 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. In an aspect of the present invention, a glutaminase preparation is presented having the recombinant glutaminase as described above wherein the preparation has low endo-protease activity. Optionally, the preparation is a liquid formulation, a dry formulation or a cell fermentate. In an aspect of the present invention, an isolated polynucleotide is presented having a nucleic acid sequence which encodes the isolated polypeptide set for the above. In another aspect of the present invention, a nucleic acid construct is presented having the isolated polynucleotide set for the above operably linked to one or more control sequences that direct the production of the polypeptide in a suitable expression host. NB41870-WO-PCT[3] In another aspect of the present invention, a recombinant expression vector is presented having the nucleic acid construct set forth above. In another aspect of the present invention, a recombinant host cell is presented having the nucleic acid construct set forth above or the vector set forth above. In another aspect of the present invention, a method for producing the recombinant glutaminase set forth above having the step of cultivating the recombinant host cell set forth above, to produce a supernatant and / or cells comprising the polypeptide; and recovering the polypeptide. In another aspect of the present invention, a recombinant glutaminase is presented which is produced by the method set forth above. BRIEF DESCRIPTION OF THE BIOLOGICAL SEQUENCES SEQ ID NO:1 shows the sequence of the AteGlt2 precursor protein. SEQ ID NO:2 shows the sequence of the AteGlt2 predicted mature protein. SEQ ID NO:3 shows the sequence of the AteGlt2 native precursor DNA. SEQ ID NO:4 shows the sequence of the EcoGlt1 precursor protein. SEQ ID NO:5 shows the sequence of the EcoGlt1 optimized precursor DNA. SEQ ID NO:6 shows the sequence of the MetGlt2 precursor protein. SEQ ID NO:7 shows the sequence of the MetGlt2 predicted mature protein. SEQ ID NO:8 shows the sequence of the MetGlt2 optimized precursor DNA. SEQ ID NO:9 shows the sequence of the MetGlt7 precursor protein. SEQ ID NO:10 shows the sequence of the MetGlt7 predicted mature protein. SEQ ID NO:11 shows the sequence of the MetGlt7 optimized precursor DNA. SEQ ID NO:12 shows the sequence of the MspGlt2 precursor protein. SEQ ID NO:13 shows the sequence of the MspGlt2 predicted mature protein. SEQ ID NO:14 shows the sequence of the MspGlt2 native precursor DNA. SEQ ID NO:15 shows the sequence of the EtaGlt1 precursor protein. SEQ ID NO:16 shows the sequence of the EtaGlt1 optimized precursor DNA. SEQ ID NO:17 shows the sequence of the TspGlt1 precursor protein. SEQ ID NO:18 shows the sequence of the TspGlt1 optimized precursor DNA. SEQ ID NO:19 shows the sequence of the SenGlt2 precursor protein. NB41870-WO-PCT[3] SEQ ID NO: 20 shows the sequence of the SenGlt2 optimized precursor DNA. SEQ ID NO:21 shows the sequence of the Amano_SD-C100S_glutaminase precursor protein. SEQ ID NO:22 shows the sequence of the subtilisin like alkaline protease precursor protein. SEQ ID NO:23 shows the sequence of the AcPepN2 N-terminal exopeptidase precursor. SEQ ID NO:24 shows the sequence of the neutral endoprotease protein. BRIEF DESCRIPTION OF THE FIGURES FIGs. 1A-1B shows formation of glutamic acid in gluten pre-hydrolysates at different temperatures FIG. 1a) reaction temperature 50˚C, FIG. 1b) reaction temperature 55˚C. All measurements were carried out in duplicates. FIG. 2 shows formation of glutamic acid, from 24 hour gluten hydrolysis at 55˚C, as a function of enzyme dose (ppm enzyme protein in reaction mixture). FIGs. 3A-3D shows specific glutaminase activity at different pH values measured at substrate (glutamine) concentrations of a) 108mM, b) 54mM, c) 27mM and d) 13,5mM. FIGs. 4A-4B shows results from enzymatic generation of glutamic acid in plant-based Frankfurter sausage. FIG. 4A) Glutamic acid levels in sausages with different enzyme treatments. The doses of enzymes are in ppm of enzyme protein relative to the total soy protein in the sample. FIG. 4B) Hardness data of the same sausages as in Figure 4A. Error bars represent STDEV (n=2). DETAILED DESCRIPTION OF THE INVENTION Definitions An “active fragment” of an enzyme is a polypeptide where the enzyme has been deleted either at the C-terminus, the N-terminus and / or internally but still retains some or all of its original activity. An active fragment includes the mature form of an enzyme. In addition to the specific amino acid sequences and polynucleotides mentioned herein, the present invention encompasses variants, homologues, derivatives, and fragments thereof. NB41870-WO-PCT[3] The protein sequences of the present invention may also have deletions, insertions or substitutions of amino acid residues which produce a silent change and result in a functionally equivalent substance. Deliberate amino acid substitutions may be made on the basis of similarity in polarity, charge, solubility, hydrophobicity, hydrophilicity, and / or the amphipathic nature of the residues as long as the secondary binding activity of the substance is retained. For example, negatively charged amino acids include aspartic acid and glutamic acid; positively charged amino acids include lysine and arginine; and amino acids with uncharged polar head groups having similar hydrophilicity values include leucine, isoleucine, valine, glycine, alanine, asparagine, glutamine, serine, threonine, phenylalanine, and tyrosine. Conservative substitutions may be made, for example according to the Table below. Amino acids in the same block in the second column and preferably in the same line in the third column may be substituted for each other as set forth in Table 1. Table 1 ALIPHATIC Non-polar G A P The present invention employs, unless otherwise indicated, conventional techniques of biochemistry, molecular biology, microbiology and recombinant DNA, which are within the capabilities of a person of ordinary skill in the art. Such techniques are explained in the literature. See, for example, J. Sambrook, E. F. Fritsch, and T. Maniatis, 1989, Molecular Cloning: A Laboratory Manual, Second Edition, Books 1-3, Cold Spring Harbor Laboratory Press; Ausubel, F. M. et al. (1995 and periodic supplements; Current Protocols in Molecular Biology, ch. 9, 13, and 16, John Wiley & Sons, New York, N. Y.); B. Roe, J. Crabtree, and A. Kahn, 1996, DNA Isolation and NB41870-WO-PCT[3] Sequencing: Essential Techniques, John Wiley & Sons; M. J. Gait (Editor), 1984, Oligonucleotide Synthesis: A Practical Approach, IrI Press; and, D. M. J. Lilley and J. E. Dahlberg, 1992, Methods of Enzymology: DNA Structure Part A: Synthesis and Physical Analysis of DNA Methods in Enzymology, Academic Press. Each of these general texts is herein incorporated by reference. As used herein, “percent (%) sequence identity” means that a particular sequence has at least a certain percentage of amino acid residues identical to those in a specified reference sequence, when aligned using the CLUSTAL W algorithm with default parameters. See Thompson et al. (1994) Nucleic Acids Res. 22:4673-4680. Default parameters for the CLUSTAL W algorithm are: Gap opening penalty: 10.0 Gap extension penalty: 0.05 Protein weight matrix: BLOSUM series DNA weight matrix: IUB Delay divergent sequences %: 40 Gap separation distance: 8 DNA transitions weight: 0.50 List hydrophilic residues: GPSNDQEKR Use negative matrix: OFF Toggle Residue specific penalties: ON Toggle hydrophilic penalties: ON Toggle end gap separation penalty: OFF Deletions are counted as non-identical residues, compared to a reference sequence. Deletions occurring at either terminus are included. For example, a variant with five amino acid deletions of the C-terminus of the mature 617 residue polypeptide would have a percent sequence identity of 99% (612 / 617 identical residues × 100, rounded to the nearest whole number) relative to the mature polypeptide. Such a variant would be encompassed by a variant having “at least 99% sequence identity” to a mature polypeptide. In accordance with the instant invention, proteins, including enzymes, of the present invention exist in multiple forms. Proteins of the instant invention may be NB41870-WO-PCT[3] clipped or trimmed (i.e., removing amino acids) from the N-terminus and / or the C- terminus, resulting in a shorter protein. Proteins of the instant invention can also have internal deletions. Shorter proteins as described herein can have higher activity or lower activity than longer counterparts. Without being bound by theory, as used herein the term precursor protein is a protein, including an enzyme, which has an N-terminal signal peptide that targets the protein for secretion. A precursor protein is sometimes referred to herein as “full length” or “full length protein”. The N-terminal signal peptide is cleaved off in the endoplasmic reticulum to yield a mature protein. A glutaminase is an amidohydrolase that converts glutamine to glutamic acid. Glutaminases may be classified in EC 3.4.1.2. However, enzymes classified as EC 2.3.2.2 (gamma glutamyl transferase) and EC 3.4.19.13 (glutathione hydrolase) may also have glutaminase activity. All references cited in the present specification are hereby incorporated by reference in their entirety. In particular, the teachings of all references herein specifically referred to are incorporated by reference. Unless otherwise defined, all terms used in disclosing the invention, including technical and scientific terms, have the meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. By means of further guidance, term definitions are included to better appreciate the teaching of the present invention. Other definitions are set forth below. Production of enzymes The enzymes of the present invention can be produced in host cells, for example, by secretion or intracellular expression. A cultured cell material (e.g., a whole-cell broth) having an enzyme can be obtained following secretion of the enzyme into the cell medium. Optionally, the enzyme can be isolated from the host cells, or even isolated from the cell broth, depending on the desired purity of the final enzyme. Suitable host cells include bacterial, fungal (including yeast and filamentous fungi), and plant cells (including algae). Particularly useful host cells include Aspergillus niger, Aspergillus oryzae or Trichoderma reesei. Other host cells include bacterial cells, e.g., Bacillus subtilis or B. licheniformis, as well as Streptomyces, E. coli. NB41870-WO-PCT[3] Vectors A DNA construct comprising a nucleic acid encoding an enzyme can be constructed to be expressed in a host cell. Because of the well-known degeneracy in the genetic code, variant polynucleotides that encode an identical amino acid sequence can be designed and made with routine skill. It is also well-known in the art to optimize codon use for a particular host cell. Nucleic acids encoding glutaminase can be incorporated into a vector. Vectors can be transferred to a host cell using well-known transformation techniques, such as those disclosed below. The vector may be any vector that can be transformed into and replicated within a host cell. For example, a vector comprising a nucleic acid encoding an enzyme can be transformed and replicated in a bacterial host cell as a means of propagating and amplifying the vector. The vector also may be transformed into an expression host, so that the encoding nucleic acids can be expressed as a functional glutaminase. Host cells that serve as expression hosts can include filamentous fungi, for example. The Fungal Genetics Stock Center (FGSC) Catalogue of Strains lists suitable vectors for expression in fungal host cells. See FGSC, Catalogue of Strains, University of Missouri, at www.fgsc.net (last modified January 17, 2007). A representative vector is pJG153, a promoterless Cre expression vector that can be replicated in a bacterial host. See Harrison et al. (June 2011) Applied Environ. Microbiol. 77: 3916-22. pJG153can be modified with routine skill to comprise and express a nucleic acid encoding a glutaminase. A nucleic acid encoding an enzyme can be operably linked to a suitable promoter, which allows transcription in the host cell. The promoter may be any DNA sequence that shows transcriptional activity in the host cell of choice and may be derived from genes encoding proteins either homologous or heterologous to the host cell. Exemplary promoters for directing the transcription of the DNA sequence encoding a glutaminase, especially in a bacterial host, are the promoter of the lac operon of E. coli, the Streptomyces coelicolor agarase gene dagA or celA promoters, the promoters of the Bacillus licheniformis α-amylase gene (amyL), the promoters of the Bacillus stearothermophilus maltogenic amylase gene (amyM), the promoters of the Bacillus amyloliquefaciens α-amylase (amyQ), the promoters of the Bacillus subtilis xylA and NB41870-WO-PCT[3] xylB genes etc. For transcription in a fungal host, examples of useful promoters are those derived from the gene encoding Aspergillus oryzae TAKA amylase, Rhizomucor miehei aspartic proteinase, Aspergillus niger neutral α-amylase, A. niger acid stable α-amylase, A. niger glucoamylase, Rhizomucor miehei lipase, A. oryzae alkaline protease, A. oryzae triose phosphate isomerase, or A. nidulans acetamidase. When a gene encoding an enzyme is expressed in a bacterial species such as E. coli, a suitable promoter can be selected, for example, from a bacteriophage promoter including a T7 promoter and a phage lambda promoter. Examples of suitable promoters for the expression in a yeast species include but are not limited to the Gal 1 and Gal 10 promoters of Saccharomyces cerevisiae and the Pichia pastoris AOX1 or AOX2 promoters. cbh1 is an endogenous, inducible promoter from Trichoderma reesei. See Liu et al. (2008) “Improved heterologous gene expression in Trichoderma reesei by cellobiohydrolase I gene (cbh1) promoter optimization,” Acta Biochim. Biophys. Sin (Shanghai) 40(2): 158-65. The coding sequence can be operably linked to a signal sequence. The DNA encoding the signal sequence may be the DNA sequence naturally associated with the enzyme gene to be expressed or from a different Genus or species. A signal sequence and a promoter sequence comprising a DNA construct or vector can be introduced into a fungal host cell and can be derived from the same source. For example, the signal sequence is the cbh1 signal sequence that is operably linked to a cbh1 promoter. An expression vector may also comprise a suitable transcription terminator and, in eukaryotes, polyadenylation sequences operably linked to the DNA sequence encoding a variant glutaminase. Termination and polyadenylation sequences may suitably be derived from the same sources as the promoter. The vector may further comprise a DNA sequence enabling the vector to replicate in the host cell. Examples of such sequences are the origins of replication of plasmids pUC19, pACYC177, pUB110, pE194, pAMB1, and pIJ702. The vector may also comprise a selectable marker, e.g., a gene the product of which complements a defect in the isolated host cell, such as the dal genes from B. subtilis or B. licheniformis, or a gene that confers antibiotic resistance such as, e.g., ampicillin, kanamycin, chloramphenicol or tetracycline resistance. Furthermore, the vector may comprise Aspergillus selection markers such as amdS, argB, niaD and xxsC, NB41870-WO-PCT[3] a marker giving rise to hygromycin resistance, or the selection may be accomplished by co-transformation, such as known in the art. See e.g., International PCT Application WO 91 / 17243. Intracellular expression may be advantageous in some respects, e.g., when using certain bacteria or fungi as host cells to produce large amounts of glutaminase for subsequent enrichment or purification. Extracellular secretion of glutaminase into the culture medium can also be used to make a cultured cell material comprising the isolated glutaminase. The expression vector typically includes the components of a cloning vector, such as, for example, an element that permits autonomous replication of the vector in the selected host organism and one or more phenotypically detectable markers for selection purposes. The expression vector normally comprises control nucleotide sequences such as a promoter, operator, ribosome binding site, translation initiation signal and optionally, a repressor gene or one or more activator genes. Additionally, the expression vector may comprise a sequence coding for an amino acid sequence capable of targeting the enzyme to a host cell organelle such as a peroxisome, or to a particular host cell compartment. Such a targeting sequence includes but is not limited to the sequence, SKL. For expression under the direction of control sequences, the nucleic acid sequence of the glutaminase is operably linked to the control sequences in proper manner with respect to expression. The procedures used to ligate the DNA construct encoding an enzyme, the promoter, terminator and other elements, respectively, and to insert them into suitable vectors containing the information necessary for replication, are well known to persons skilled in the art (see, e.g., Sambrook et al., MOLECULAR CLONING: A LABORATORY MANUAL, 2nded., Cold Spring Harbor, 1989, and 3rded., 2001). Transformation and Culture of Host Cells An isolated cell, either comprising a DNA construct or an expression vector, is advantageously used as a host cell in the recombinant production of an enzyme according to the instant invention. The cell may be transformed with the DNA construct encoding the enzyme, conveniently by integrating the DNA construct (in one or more copies) in the host chromosome. This integration is generally considered to be an advantage, as the NB41870-WO-PCT[3] DNA sequence is more likely to be stably maintained in the cell. Integration of the DNA constructs into the host chromosome may be performed according to conventional methods, e.g., by homologous or heterologous recombination. Alternatively, the cell may be transformed with an expression vector as described above in connection with the different types of host cells. Examples of suitable bacterial host organisms are Gram positive bacterial species such as Bacillaceae including Bacillus subtilis, Bacillus licheniformis, Bacillus lentus, Bacillus brevis, Geobacillus (formerly Bacillus) stearothermophilus, Bacillus alkalophilus, Bacillus amyloliquefaciens, Bacillus coagulans, Bacillus lautus, Bacillus megaterium, and Bacillus thuringiensis; Streptomyces species such as Streptomyces murinus; lactic acid bacterial species including Lactococcus sp. such as Lactococcus lactis; Lactobacillus sp. including Lactobacillus reuteri; Leuconostoc sp.; Pediococcus sp.; and Streptococcus sp. Alternatively, strains of a Gram negative bacterial species belonging to Enterobacteriaceae including E. coli, or to Pseudomonadaceae can be selected as the host organism. A suitable yeast host organism can be selected from the biotechnologically relevant yeasts species such as but not limited to yeast species such as Pichia sp., Hansenula sp., or Kluyveromyces, Yarrowinia, Schizosaccharomyces species or a species of Saccharomyces, including Saccharomyces cerevisiae or a species belonging to Schizosaccharomyces such as, for example, S. pombe species. A strain of the methylotrophic yeast species, Pichia pastoris, can be used as the host organism. Alternatively, the host organism can be a Hansenula species. Suitable host organisms among filamentous fungi include species of Aspergillus, e.g., Aspergillus niger, Aspergillus oryzae, Aspergillus tubigensis, Aspergillus awamori, or Aspergillus nidulans. Alternatively, strains of a Fusarium species, e.g., Fusarium oxysporum or of a Rhizomucor species such as Rhizomucor miehei can be used as the host organism. Other suitable strains include Thermomyces and Mucor species. In addition, Trichoderma sp. can be used as a host. A suitable procedure for transformation of Aspergillus host cells includes, for example, that described in EP 238023. An enzyme expressed by a fungal host cell can be glycosylated, i.e., will comprise a glycosyl moiety. The glycosylation pattern can be the same or different as present in the wild-type glutaminase. The type NB41870-WO-PCT[3] and / or degree of glycosylation may impart changes in enzymatic and / or biochemical properties. It may be advantageous to delete genes from expression hosts, where the gene deficiency can be cured by the transformed expression vector. Known methods may be used to obtain a fungal host cell having one or more inactivated genes. Gene inactivation may be accomplished by complete or partial deletion, by insertional inactivation or by any other means that renders a gene nonfunctional for its intended purpose, such that the gene is prevented from expression of a functional protein. Any gene from a Trichoderma sp. or other filamentous fungal host that has been cloned can be deleted, for example, cbh1, cbh2, egl1, and egl2 genes. Gene deletion may be accomplished by inserting a form of the desired gene to be inactivated into a plasmid by methods known in the art. Introduction of a DNA construct or vector into a host cell includes techniques such as transformation; electroporation; nuclear microinjection; transduction; transfection, e.g., lipofection mediated and DEAE-Dextrin mediated transfection; incubation with calcium phosphate DNA precipitate; high velocity bombardment with DNA-coated microprojectiles; and protoplast fusion. General transformation techniques are known in the art. See, e.g., Sambrook et al. (2001), supra. The expression of heterologous protein in Trichoderma is described, for example, in U.S. Patent No. 6,022,725. Reference is also made to Cao et al. (2000) Science 9:991-1001 for transformation of Aspergillus strains. Genetically stable transformants can be constructed with vector systems whereby the nucleic acid encoding an enzyme is stably integrated into a host cell chromosome. Transformants are then selected and purified by known techniques. Expression A method of producing an enzyme of the instant invention may comprise cultivating a host cell as described above under conditions conducive to the production of the enzyme and recovering the enzyme from the cells and / or culture medium. The medium used to cultivate the cells may be any conventional medium suitable for growing the host cell in question and obtaining expression of the enzyme. Suitable media and media components are available from commercial suppliers or may be prepared according to published recipes (e.g., as described in catalogues of the American NB41870-WO-PCT[3] Type Culture Collection). An enzyme secreted from the host cells can be used in a whole broth preparation. In the present methods, the preparation of a spent whole fermentation broth of a recombinant microorganism can be achieved using any cultivation method known in the art resulting in the expression of a glutaminase. Fermentation may, therefore, be understood as comprising shake flask cultivation, small- or large-scale fermentation (including continuous, batch, fed-batch, or solid-state fermentations) in laboratory or industrial fermenters performed in a suitable medium and under conditions allowing the glutaminase to be expressed or isolated. The term “spent whole fermentation broth” is defined herein as unfractionated contents of fermentation material that includes culture medium, extracellular proteins (e.g., enzymes), and cellular biomass. It is understood that the term “spent whole fermentation broth” also encompasses cellular biomass that has been lysed or permeabilized using methods well known in the art. An enzyme secreted from the host cells may conveniently be recovered from the culture medium by well-known procedures, including separating the cells from the medium by centrifugation or filtration, and precipitating proteinaceous components of the medium by means of a salt such as ammonium sulfate, followed by the use of chromatographic procedures such as ion exchange chromatography, affinity chromatography, or the like. Host cells may be cultured under suitable conditions that allow expression of a glutaminase. Expression of the enzymes may be constitutive such that they are continually produced, or inducible, requiring a stimulus to initiate expression. In the case of inducible expression, protein production can be initiated when required by, for example, addition of an inducer substance to the culture medium, for example dexamethasone or IPTG or Sophorose. Polypeptides can also be produced recombinantly in an in vitro cell-free system, such as the TNT™ (Promega) rabbit reticulocyte system. Methods for Enriching and Purifying enzymes Fermentation, separation, and concentration techniques are well known in the art and conventional methods can be used in order to prepare an enzyme polypeptide- containing solution. After fermentation, a fermentation broth is obtained, the microbial cells and NB41870-WO-PCT[3] various suspended solids, including residual raw fermentation materials, are removed by conventional separation techniques in order to obtain an enzyme solution. Filtration, centrifugation, microfiltration, rotary vacuum drum filtration, ultrafiltration, centrifugation followed by ultra-filtration, extraction, or chromatography, or the like, are generally used. It is desirable to concentrate an enzyme polypeptide-containing solution in order to optimize recovery. Use of unconcentrated solutions requires increased incubation time in order to collect the enriched or purified enzyme precipitate. The enzyme containing solution is concentrated using conventional concentration techniques until the desired enzyme level is obtained. Concentration of the enzyme containing solution may be achieved by any of the techniques discussed herein. Exemplary methods of enrichment and purification include but are not limited to rotary vacuum filtration and / or ultrafiltration. Description of the Preferred Embodiments In an aspect of the present invention, a method for producing a hydrolysate enriched in glutamate is presented having the step of adding a glutaminase corresponding to an isolated polypeptide having at least 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof to a starting material comprising glutamine and incubating for a sufficient time to produce the hydrolysate. Preferably, the isolated polypeptide has at least 80% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. More preferably, the isolated polypeptide has at least 85% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Yet more preferably, the isolated polypeptide has at least 90% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Still more preferably, the isolated polypeptide has at least 95% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NB41870-WO-PCT[3] NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. In still more preferred embodiments, the isolated polypeptide has at least 98% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. More preferably, the isolated polypeptide has at least 99% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Most preferably, the isolated polypeptide has 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Preferably, the step of incubating is conducted at a temperature of around 40 to 70°C. More preferably, the step of incubating is conducted at a temperature of around 45 to 65°C. Still more preferably, the step of incubating is conducted at a temperature around 50 to 60°C. Most preferably, the step of incubating is conducted at a temperature around 55°C. Preferably, the starting material is a plant protein. More preferably, the plant protein is selected from the group consisting of pea, soy, fava, gluten, and oat. Preferably, the method has the further step of adding an exo-peptidase. Preferably, the exo-peptidase is an isolated polypeptide having at least 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:23 or an exopeptidase active fragment thereof. Preferably, the method has the further step of adding an endo-peptidase. Preferably, the endopeptidase is an isolated polypeptide having at least 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:22 or an endopeptidase active fragment thereof or SEQ ID NO:24 or an endopeptidase active fragment thereof. In another aspect of the present invention, a protein hydrolysate enriched in glutamate is presented which is produced according to the method described above. In another aspect of the present invention, use of a protein as made above to produce a broth, a meat product or a cheese is presented. In another aspect of the present invention, a method of providing umami flavor to a plant-based meat alternative is presented having the steps of: NB41870-WO-PCT[3] a. providing a batter comprising a plant protein; b. combining the batter with a glutaminase enzyme comprising an isolated polypeptide having at least 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof; c. incubating the batter at a temperature of about 40 to 70°C; and d. cooking the batter to provide the plant-based meat alternative. Preferably, the isolated polypeptide has at least 80% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. More preferably, the isolated polypeptide has at least 85% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Yet more preferably, the isolated polypeptide has at least 90% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Still more preferably, the isolated polypeptide has at least 95% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. In still more preferred embodiments, the isolated polypeptide has at least 98% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. More preferably, the isolated polypeptide has at least 99% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Most preferably, the isolated polypeptide has 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Preferably, the step of incubating is conducted at a temperature of around 40 to 70°C. More preferably, the step of incubating is conducted at a temperature of around 45 to 65°C. Still more preferably, the step of incubating is conducted at a temperature NB41870-WO-PCT[3] around 50 to 60°C. Most preferably, the step of incubating is conducted at a temperature around 55°C. Preferably, the plant protein is selected from the group consisting of pea, soy, fava, gluten, and oat. Preferably, has the further step of adding an exo-peptidase at step a and / or step b. Preferably, the exo-peptidase is an isolated polypeptide having 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:23 or a proteolytically active fragment thereof. Preferably, the method has the further step of adding an endo-peptidase at step a and / or step b. Preferably, the endo-peptidase comprises an isolated polypeptide having 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:22 or SEQ ID NO:24 or an exopeptidase active fragment thereof. In another aspect of the present invention, a recombinant glutaminase is presented which is an isolated polypeptide having at least 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof to a starting material comprising glutamine and incubating for a sufficient time to produce the hydrolysate. Preferably, the isolated polypeptide has at least 80% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. More preferably, the isolated polypeptide has at least 85% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Yet more preferably, the isolated polypeptide has at least 90% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Still more preferably, the isolated polypeptide has at least 95% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. In still more preferred embodiments, the isolated polypeptide has at least 98% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, NB41870-WO-PCT[3] SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. More preferably, the isolated polypeptide has at least 99% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. Most preferably, the isolated polypeptide has 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof. In an aspect of the present invention, a glutaminase preparation is presented having the recombinant glutaminase as described above wherein the preparation has low endo-protease activity. Preferably, the preparation is a liquid formulation, a dry formulation or a cell fermentate. In an aspect of the present invention . an isolated polynucleotide is presented having a nucleic acid sequence which encodes the isolated polypeptide set for the above. In another aspect of the present invention, a nucleic acid construct is presented having the isolated polynucleotide set for the above operably linked to one or more control sequences that direct the production of the polypeptide in a suitable expression host. In another aspect of the present invention, a recombinant expression vector is presented having the nucleic acid construct set forth above. In another aspect of the present invention, a recombinant host cell is presented having the nucleic acid construct set forth above or the vector set forth above. In another aspect of the present invention, a method for producing the recombinant glutaminase set forth above having the step of cultivating the recombinant host cell set forth above, to produce a supernatant and / or cells comprising the polypeptide; and recovering the polypeptide. In another aspect of the present invention, a recombinant glutaminase is presented which is produced by the method set forth above. The present disclosure is described in further detail in the following examples, which are not in any way intended to limit the scope of the disclosure as claimed. The attached figures are meant to be considered as integral parts of the specification and NB41870-WO-PCT[3] description of the disclosure. The following examples are offered to illustrate, but not to limit the claimed disclosure. EXAMPLES Example 1. Cloning and transformation of glutaminases The glutaminase genes were identified either from a public database (JGI or NCBI) or from internal genome databases, and the nucleotide and corresponding polypeptide sequences are shown on Table 1. For the bacterial glutaminases, genes encoding the predicted mature polypeptide region were codon-optimized based on Bacillus subtilis codon preference and cloned into the p2JM vector (Vogtentanz, Protein Expr Purif. 55:40-52, 2007). The AprE promoter and signal peptide were used for transcription and secretion. The constructed vector was subjected to rolling-circle amplification and transformed into the host B. subtilis CBS6. Generally, the fungal glutaminase genes were synthesized and cloned into a vector (same as the pTTTpyr2 vector as described in published PCT Application WO2015 / 017256, incorporated by reference herein) by Generay Biotech (Shanghai, China). The constructed vectors were transformed into a suitable Trichoderma reesei host strain (described in published PCT application WO 05 / 001036) using protoplast transformation (Te’o et al. (2002) J. Microbiol. Methods 51:393-99). Table 1. Sequence list of glutaminases Gene name Expressed in Description SEQ ID No. NB41870-WO-PCT[3] MspGlt2 Trichoderma reesei Precursor PRT 12 Predicted Mature PRT 13 To initiate T. reesei fermentation, seed culture was grown in 1 L shake flask, each contained 100 mL defined medium (pH 5.5 before sterilization) containing 25 g / L glucose, 6 g / L glycine, 5 g / L (NH4)2SO4, 4.5 g / L KH2PO4, 1 g / L CaCl2·2H2O, 1 g / L MgSO4·7H2O, 2 g / L Mazu 6000K and 2.5 mL 400× T. reesei Trace Metals (175 g / L C6H8O7·H2O, 200 g / L FeSO4·7H2O, 16 g / L ZnSO4·7H2O, 3.2 g / L CuSO4·5H2O, 1.4 g / L MnSO4·H2O, and 0.8 g / L H3BO3). The seed culture was shaken for 48 hours at 250 rpm and 30°C. Then, 200 mL seed culture was transferred to the 2-L bioreactor (DASGIP) containing 1 L medium (50 g / L glucose, 6 g / L glycine, 1 g / L CaCl2·2H2O, 4.5 g / L KH2PO4, 4 g / L (NH4)2SO4, 1 g / L MgSO4·7H2O, 1.2 g / L Mazu 6000K and 2.5 ml 400× T. reesei Trace Metals). Following inoculation, batch fermentation was initiated and was controlled at pH 3.5 and 34 °C. Dissolved oxygen level was controlled above 30% by adjusting agitation, airflow rate and oxygen supply during whole fermentation process. After 18 hours, the glucose in broth was depleted, and the feed of 250 g (glucose / sophorose) / kg solution was started. Stepwise feed rates of 4 mL feed / hr and 6 mL feed / h were applied in the time intervals of 22-46 hours and 46-72 hours, respectively. Accompanying the start of fed-batch phase, pH was changed to 4.0, and temperature was adjusted to 28 °C. Fermentation was completed after 72 hours run, and culture supernatant was harvested by centrifugation, filtered, and subsequently concentrated. The B. subtilis CBS6 transformants were streaked on LB plate supplementing 5 ppm chloramphenicol. The colony was inoculated into 20 ml LB medium with 5 ppm chloramphenicol in 250 ml shake flask for seed growth. Well-grown cells were inoculated into 350 ml Grant’s II medium supplementing 5 ppm chloramphenicol in a 3- NB41870-WO-PCT[3] L ultrayield shake flask and incubate at 37°C with vigorous shaking at 250 rpm overnight and then 32°C for 24 h. Culture supernatant was harvested by centrifugation, filtered, and subsequently concentrated. Example 3. Purification of glutaminases Ammonium sulfate was added to the concentrated fermentation broth of T. reesei to a final concentration of 1 M. The solution was loaded onto HiPrepTMPhenyl FF 16 / 10 column pre-equilibrated with 20 mM NaAc (pH 5.0) containing 1 M ammonium sulfate, then eluted with 20 mM NaAc (pH 5.0) buffer in a stepwise mode of decreasing ammonium sulfate concentration. The fractions containing target protein were combined, buffer-exchanged to 20 mM NaAc (pH 5.0) and loaded onto HiPrepTMQ 16 / 10. The target protein was eluted through a gradient of 0-1 M NaCl in 20 mM NaAc (pH 5.0) buffer. The fractions containing target protein were combined, concentrated and buffer- exchanged to 20 mM NaAc (pH 5.0) 150 mM NaCl in Amicon Ultra ultrafiltration tubes (10 kD cutoff) and stored in 40% glycerol at -20 °C until usage. Purification of glutaminase in B. subtilis fermentation broth is similar as T. reesei, except that 20 mM Tris-HCl (pH 7.5) buffer system was used instead of NaAc. Example 4. Specific activity of glutaminases The experimental procedure in this example was based on glutamine deamidation by glutaminase and measurement of the released ammonium. All reagents were of analytical grade or purer. L-Glutamine, and N-2-Hydroxyethylpiperazine-Nʹ-2-ethanesulfonic acid (HEPES) were from Merck (Soeborg DK). A working solution with a HEPES of 50 mM and a glutamine concentration of 10 mM was obtained by dissolving 2.38 g of HEPES and 0.29 g glutamine in 200 mL ultrapure water. For assay purposes, enzymes were serial diluted in ultrapure water to give final responses within the dynamic range of the assay. To start the reaction, 10 µL of the diluted enzyme or water (blank) was mixed with 90 µL of the glutamine substrate solutions in a 96-MTP (Corning 3641). The plate was sealed and incubated in a Thermomixer at 50 °C and 650 rpm for 10 min. At the end of incubation, free ammonium was quantified by spectrofluorimetric method using OPA / NAC reagent. The OPA / NAC reagent was prepared freshly by mixing together 1 volume of reagent A (200 mM o-phthalaldehyde in ethanol), 1 volume of reagent B (200 mM N-acetyl-L-cysteine and 20 mM tris(2- NB41870-WO-PCT[3] carboxyethyl) phosphine hydrochloride in 1 M HCl) and 18 volumes of buffer (0.1 M sodium borate buffer, pH 10.5).5 μL of sample or standard solution (ammonium chloride) were transferred into a 96-well microtiter plate (Corning 3605) containing 50 μL of OPA / NAC reagent. The plate was shaken for 5 min and fluorescence intensity was then recorded with excitation and emission wavelengths set at λex = 415 nm / λem = 485 nm using a Microplate reader (Molecular devices, SpectraMax iD3). Concentrations in samples were determined using the linear calibration curves obtained with standards. The ammonium amount in the substrate was subtracted from the amount of each reaction mixture. One unit of activity was defined as the amount of enzyme required to catalyze the release of one µmole of ammonia per minute under the described conditions. The specific activity is summarized in Table 2. The deamidating activity results using L-glutamine as the substrate indicated that AteGlt2, EcoGlt1, MetGlt2, MetGlt7, MspGlt2, EtaGlt1, TspGlt1, and SenGlt2 were all active glutaminases. The Amano SD-C100S Glutaminase (Amano Enzyme U.S.A. Co., Ltd.) was used as a control and is the current market standard for glutaminases (hereinafter “Amano enzyme”). The data shown in Table 2 indicate that TspGlt1 is the most active enzyme, followed by EcoGlt1, EtaGlt1, and AteGlt2, which were more active than the Amano enzyme. This indicates that TspGlt1, EcoGlt1, EtaGlt1, and AteGlt2 may be more efficient enzyme samples to use than the current market standard. Table 2. Specific activity (U / mg) Amano AteGlt2 EcoGlt1 MetGlt2 MetGlt7 MspGlt2 EtaGlt1 TspGlt1 SenGlt2 me Example 5. pH profile of glutaminases The pH profile of AteGlt2, EcoGlt1, MetGlt2, MetGlt7, MspGlt2, EtaGlt1, TspGlt1, SenGlt2 and Amano enzyme was studied with L-glutamine as the substrate, in 25 mM NaAc / Glycine / HEPES buffer with different pH values ranging from 3 to 10. To initiate the assay, 45 μL of 50 mM NaAc / Glycine / HEPES buffer with a specific pH was first mixed with 45 μL of 20 mM L-glutamine in a 96-MTP and pre-incubated at 50 °C for 5 min, NB41870-WO-PCT[3] followed by the addition of 10 μL of water diluted enzyme or water (the blank control). The reaction was performed and analyzed as described in Example 4. Enzyme activity at each pH was reported as the relatively activity, where the activity at the optimal pH was set to be 100%. The pH range tested was 3, to 10 as shown on Table 3. Table 3. pH profile Relative activity (%) pH Amano e It was determined that AteGlt2, MetGlt2, MetGlt7, MspGlt2, and Amano enzyme preferred alkaline conditions. The EcoGlt1, EtaGlt1, TspGlt1, and SenGlt2 enzymes preferred acidic conditions. Example 6. Temperature activity profile of glutaminases The temperature activity profile was analyzed in 50 mM HEPES (pH 7) using L- glutamine as the substrate for AteGlt2, MetGlt2, MetGlt7, and MspGlt2 enzymes. The temperature profile was analyzed in 50 mM sodium acetate (pH 5) using L-glutamine as the substrate for EcoGlt1, EtaGlt1, TspGlt1, and SenGlt2. The enzyme sample and L- glutamine substrate were prepared as in Example 4. Prior to the reaction, 90 μL of L- glutamine dissolved in 50 mM HEPES (pH 7) or sodium acetate (pH 5) was transferred in a 200 μL PCR tube, which was then incubated in a Peltier Thermal Cycler (BioRad) at desired temperatures (i.e.30~80 °C) for 5 min. After the incubation, 10 μL of water diluted enzyme or water (the blank control) was added to the solution, and the reaction was carried out in the Peltier Thermal Cycle for 10 min at different temperatures. The reaction was performed and analyzed as described in Example 4. The activity was reported as the relative activity, where the activity at the optimal temperature was set to be 100%. NB41870-WO-PCT[3] Table 4. Temperature activity profile at pH 7 Relative activity (%) Temp (℃) At Glt2 M tGlt2 M tGlt7 M Glt2 Amano ratures for AteGlt2, MetGlt2, MetGlt7, MspGlt2, and Amano enzymes activity were 53, 60, 50, 55, and 53 °C, respectively. As shown in Table 5, the optimal temperatures when EcoGlt1, EtaGlt1, TspGlt1, SenGlt2, and Amano enzyme were tested at pH 5, were 60, 53, 50, 50, and 50 °C, respectively. Table 5. Temperature profile at pH 5 Relative activity (%) ° 3 2 2 8 0 5 3 5 9 0 0 NB41870-WO-PCT[3] 80 12 6 6 0 0 buffer, with L-glutamine as the substrate, and with pre-incubated at different pH conditions: 20 mM of sodium citrate buffer (pH 3, and pH 4), 20 mM of sodium acetate buffer (pH 5), 20 mM of sodium phosphate buffer (pH 6, and pH 7), and 20 mM of HEPES buffer (pH 7) at room temperature for 1 hour incubation before measuring the remaining activity. The activity of enzyme samples pre-incubated in 20 mM HEPES (pH 7) at 4oC was set to be 100%. Prior to the pre-incubation, the enzyme was diluted to 1 mg / mL with 40% (w / w) glycerol. To initiate the pre-incubation, 90 μL of buffer mentioned was mixed with 10 μL of enzyme samples and in a 96-MTP; and after 60 min pre-incubation at room temperature, dilute enzyme sample with water to a concentration that gives response of excitation and emission wavelengths set at λex = 415 nm / λem = 485 nm within the linear part of glutaminase reaction curve. The reaction was performed and analyzed as described in Example 4. Table 6. Enzyme stability at room temperature for 1 hour at various pH (pH 3 to 7). Relative activity (%) H o e s s own n ab e 6, w en es ed a p 3, eG , coG , spG , aG , spG and SenGlt2 retain >50% of their activity after 1-hour pre-incubation, with TspGlt1 and SenGlt2, retaining >90% of their activity, while the Amano enzyme only retained 7% of its activity. Example 8. Product inhibition of glutaminases Product (glutamic acid) inhibition was investigated using 50 mM HEPES (pH 7) as the incubation buffer, with L-glutamine as the substrate, with addition of glutamic acid NB41870-WO-PCT[3] concentrations of 0.16, 0.31, 0.63, 1.25, 2.5, and 5 mM for determination of glutaminase activity. Prior to the reaction, the enzymes were diluted with water to specific concentrations to normalize the activity across the samples. To initiate the assay, 90 μL of L-glutamine solution with 0, 0.16, 0.31, 0.63, 1.25, 2.5, or 5 mM glutamic acid was mixed with 10 μL of the diluted enzyme samples (or water as the blank) in a 96-MTP; and after 10 min incubation at 50oC in a Thermomixer, the released ammonium was measured as described in Example 4. The enzyme activity was reported as the relative activity compared to the activity without the addition of glutamic acid set to 100%. Table 7. Measurement of relative product inhibition, by glutamic acid Gluta Relative activity (%) mic o e As shown in Table 7, EcoGlt2, MspGlt2, EtaGlt1, TspGlt1, and SenGlt2 showed significantly less product inhibition compared to the Amano enzyme. The product inhibition of AteGlt2, MetGlt2 and MetGlt7 were on par with Amano enzyme. Example 9. Glutamic acid synthesis from wheat gluten Gluten pre-hydrolysate was made using a modified version of the method described in Schlichtherle-Cerny and Amadò (2002). Following was mixed in a 100 mL screw cap bottle: 10.7 g Gluten (TCI, Japan), 93 µL FoodPro® Alkaline Protease (DuPont® Industrial Biosciences, Brabrand Denmark) (SEQ ID NO:22) (hereinafter “alkaline protease”), 12 mg aminopeptidase protein (AcPepN2, obtained according to WO2016210395 A1) (SEQ ID NO:23) and 2.9 g NaCl (Analytical grade, Fischer Scientific, Roskilde, Denmark). The total content was made up to 52.5 g with Milli-Q water. The bottle was incubated for 16 hours at 55˚C in a shaker (INFOS HT, NB41870-WO-PCT[3] Switzerland) at 250 rpm. Subsequently enzymes were inactivated by heating in a water bath kettle (Thermo Fisher scientific, USA) at 95˚C for 15 min, the slurry was centrifuged for 20 min at 10,000 rpm and the supernatant was collected and filtered through 0.45 µm vacuum filters to make the final gluten pre-hydrolysate. Glutamic acid synthesis was studied with gluten pre-hydrolysate as the substrate. 90 µL / well of gluten pre-hydrolysate, and 10 µL / well glutaminase or water was mixed in 96- MTP and sealed with thermo sealer (Biosero, Chameleon XT). All samples were run at 0.63, 1.25, 2.5 and 5 ppm and in triplicate at 50, and 55 °C. Incubation time was 16 h in a Thermomixer (Eppendorf, Comfort 5355) at 800 rpm. After incubation 30 µL / well of 30% (w / w) TCA was added to stop the reaction. Glutamic acid was determined in supernatants from the stopped reaction mixture. Glutamic acid was quantified using Enzymatic L-glutamic acid kit from Sangon Biotech. The 5 components of the kit were treated according to manufacturer’s prescriptions resulting in Solutions 1, Solution 2, and Solution standard. The method described in the kit was modified and downscaled for use in 96-well plates. Solution 1 was made by mixing 55 mL of buffer 1 with reagent 3. Solution 2 was made by mixing 3.5 mL of buffer 2 with reagent 4. Standards were made as a series of 1:1 dilutions of Solution Standard. The following was mixed in wells of a 96-well plate: 160 µL of Solution 1, 10 µL of Solution 2 and 10 µL sample, standard or water (blank). The components were mixed by shaking with Vortex-microplate genie (Scientific industries, USA). The plate was placed in a SpectraMax ABS Plus (Molecular Devices, USA) and absorbance was measured at 340 nm every 30 sec for 5 min. The increasement of readings from 0 to 5 min was used to calculate glutamic acid amount from the standard curve. The results of the gluten pre-hydrolysate experiment were shown in Table 8 and Table 9. Glutamic acid yield was depicted as a function of glutaminase protein dose at 0.63, 1.25, 2.5 and 5 ppm. At 50 and 55 °C, EcoGlt1, EtaGlt1, and TspGlt1 obviously outperformed Amano enzyme and other glutaminases. The performance of AteGlt2, MetGlt2, and MetGlt7were on par with Amano enzyme at 50 and 55°C. SenGlt2 showed significantly better performance at 50 °C but poorer performance at 55°C than Amano enzyme. NB41870-WO-PCT[3] Table 8. Gluten pre-hydrolysate activity at 50 °C for 16 hours Released glutamic acid amount (mM) Enzym e (ppm) AteGlt EcoGlt MetGlt MetGlt MspGlt TspGlt SenGlt Amano Et Glt1 e hoursReleased glutamic acid amount (mM) Enzyme Amano e Gluten pre-hydrolysate, glutaminase and water was mixed in Eppendorf tubes according to Table 6. Purified AteGlt2 enzyme with a protein concentration of 6.6 mg / mL was used, and a sample of Amano enzyme with a protein concentration of 3 mg / mL was made and used (protein concentration in the Amano enzyme sample was determined using a rapid MAX N protein analyzer from Elemantar, Langenselbold DE). All samples were run in duplicate at 50, and 55 °C. Incubation time was 14h in a Thermomixer at 1000 rpm. After incubation 0.275mL 2.5M TCA was added to stop the reaction. Glutamic acid was determined in supernatants from the stopped reaction mix. according to Example 9. Table 10: Experimental setup for investigating glutamine conversion by glutaminase in gluten pre-hydrolysates at different temperatures. NB41870-WO-PCT[3] 6 1 - 20 88 54.2 7 1 - 40 68 108.3 -1B. Glutamic acid yield is depicted as a function of glutaminase protein dose. At 55 °C, performance of AteGlt2 lies above the performance of Amano enzyme for all substrate concentrations tested (Figure 1B). At 50 °C, performance of AteGlt2 also lies above the performance of Amano enzyme at enzyme doses up to around 100 ppm. At 168 ppm (the highest dose tested for AtGlt2) performance in terms of glutamic acid yield is roughly on par with Amano enzyme (Figure 1A). The results show that under most conditions that are relevant for gluten hydrolysis AteGlt2 enzyme performs better than the Amano enzyme. For prolonged hydrolysis it is valuable to increase the temperature from 50 to 55˚C since it will increase reaction rate (reduce incubation time) and protect better against microbial spoilage. Example 11. Gluten Hydrolysis The gluten hydrolysis setup was modified from a procedure described previously (Schlichtherle-Cerny and Amadò 2002). But due to the results obtained in Example 11, incubation temperature was increased to 55˚C. In 20 mL Wheaton vials 1 g gluten powder (containing 85% gluten protein), 0.2595 g NaCl and enzymes were mixed and made up to 5g with cold tap water. The enzymes used were: alkaline protease, which was added at a dose of 1% relative to gluten, AcPepN2, which was added at a dose of 1.3 mg enzyme protein pr. g. gluten and glutaminase which was added at doses between 1,4 and 73 µg enzyme protein pr. gram gluten (corresponding to between 6 and 325 ppm enzyme protein relative to the reaction mix). The vials were placed in a heating block at 55 °C with stirring (450rpm). After 1 hour, pH was adjusted to 6.5 with 1M NaOH or 1M HCL. Reaction was stopped after 24 hours of hydrolysis by adding 1.92mL 1.8M TCA. Glutamic acid concentration was determined, in supernatants from the stopped reaction mix. using the glutamic acid assay described in Example 9. The results of the gluten hydrolysis experiments are shown in Figure 2. Glutamic acid yield is depicted as a function of glutaminase dose. It is seen that AteGlt2 performs markedly better than Amano enzyme at all substrate concentrations. NB41870-WO-PCT[3] Example 12. Ammonia determination using OPA reagent The assay is based on ammonia determination and modified from a procedure previously described (Robert-Peillard, Barco et al. 2017). All reagents used were of analytical grade or purer. N-Acetyl-L-Cysteine (NAC), Tris(2-carboxyethyl) phosphine hydrochloride (TCEP), o-Phthaldialdehyde (OPA) and sodium tetraborate decahydrate were from Merck (Soeborg DK). Hydrochloric acid 37% and Ammonium sulphate were from Fisher Scientific (Roskilde DK). Ethanol, 99.5% was from VWR (Soeborg DK). Stock solutions of 100 mM Ammonium sulphate, 1 M HCl and 0.1 M Sodium Borate buffer pH 10.5 were prepared in ultrapure water. Reagents were prepared fresh before assay: Reagent A: 200 mM NAC and 15 mM TCEP in 1M HCl and reagent B: 130 mM OPA in ethanol. Assay reagent was prepared by mixing Reagent A, Reagent B and Sodium Borate buffer in the ratio 6:4:90. Ammonium sulphate solution was diluted to make standards in the range 2 – 100 mM. For the assay, 10 µl sample, standards or blank was mixed with 100 µl assay reagent in a 96-well fluorescence compatible plate (Costar 3915, Corning Inc. Kennebunk ME, USA). The plate was sealed and incubate 10 min at 30 °C and 300 rpm in an Eppendorf Thermomixer. Immediately after incubation the seal was removed, the plate was placed in a spectrofluorometer (Spectra Max Gemini EM, Molecular Devices, San Jose CA, USA) and top fluorescence was measured using λex415 nm and λem485 nm. The fluorescence values were used to calculate ammonia concentration, from a standard curve made on basis of the ammonium sulphate standards. Example 13. Investigation of specific activities The experimental procedure in this example is based on glutamine deamidation by glutaminase and measurement of the released ammonia. All reagents were of analytical grade or purer. L-Glutamine, L-glutamic acid and Citric acid monohydrate (Fluka) were from Merck (Soeborg DK). A stock solution with a citrate concentration of 70 mM and a glutamine concentration of 144mM was obtained by dissolving 3,35 g of citric acid and 5,25 g glutamine in approximately 200 mL ultrapure water. The pH was adjusted to 4 with 5M NaOH and the total volume was brought to 250 mL with ultrapure water. Citrate-glutamine solutions at different pH values ranging from 4 to 9 were obtained by placing 7,5mL aliquots of the stock solution in tarred Wheaton vials, adjusting pH with 1M NaOH and bringing the total weight of solutions to 10 g by adding Ultrapure water. NB41870-WO-PCT[3] This resulted in a solution consisting of 108 mM glutamine, 52 mM citrate solutions at different pH values. These solutions were further diluted with ultrapure water to give a range of substrate (glutamine) concentrations at different pH values. For assay purposes enzymes were diluted in ultrapure water to give final responses within the dynamic range of the assay. To start the reaction, 10 µl of the diluted enzyme was mixed with 90 µL of the glutamine-citrate substrate solutions in a 96 well plate (Thermo Scientific 269620, Roskilde DK). The plate was sealed and incubate in a Thermomixer at 50 °C and 300rpm for 30 min. Immediately after the incubation, enzyme-substrate solutions were diluted x10 in Ultrapure water and analyzed using the procedure described in Example 12. All experiments were performed in duplicate. Results from the experiments are shown in Figures 3A-3D. It is seen that at all substrate concentrations and pH values tested, AteGlt2 has a higher specific activity than Amano enzyme. Example 14. Use of glutaminase in plant-based meat alternative preparation Wet ingredients used in the recipe are: 600 g tap water, 270 g ice and 64.8g rapeseed oil. Dry ingredients are: 132g powdered soy protein isolate (Supro® EX 45), 120 g extruded soy protein (Supro® Max 5013 IP), 6g carrageenan (GRINSTED® Carrageenan 100 from IFF inc.), 3g potassium chloride, 12.2g salt, 3.6g onion powder, 0.9g ground pepper, 1.4g paprika and 1.8g glucose. Initially all wet ingredients were added to a Robot Coupe R602V blender and mixed at high speed for 20 sec to make an emulsion. All dry ingredients, except extruded protein, were mixed, and blended into the emulsion at medium speed for 20 sec. Extruded protein was added and mixed into the batter by a 10 sec blend at low speed. The final batter was left to hydrate for 1h. After hydration the batter was blended 2 times 1.5 min. In between blends the batter was scraped down from the sides of the container. The batter was further mixed by hand around 2 min and then divided into portions of 70 g. Enzymes were mixed into these portions by thorough hand mixing. The batters were then stuffed into 50 mL centrifuge tubes and centrifuged at 4000 rpm for 5min (to remove air bubbles). The tubes were closed and placed in a 50 ˚C water bath for 1h, transferred to a 90 ˚C water bath where they incubated for 20 min. After this cook cycle the tubes were placed in plenty of cold water until the temperature had reached room temperature. The resulting plant-based sausages were removed from NB41870-WO-PCT[3] the tubes and cut in slices of 2,5 cm. Texture profile analysis (TPA) was carried out on the sausage using a TA.XTPlusC Texture Analyser – 650H from Stable Microsystems. For glutamic acid measurement, 2,0 g of sausage was dispersed in 20 g water. The slurry was incubated at room temperature for 1 hour with magnetic stirring (240 rpm). Approximately 1 mL of supernatant was filtered through a 0,2 µm syringe filter. Glutamic acid was measured on the permeate according to the method described in Example 9. The enzymes used in this study were AteGlt2 (SEQ ID NO: 2), AcPepN2 (SEQ ID NO:23) and FoodPro® PNL (DuPont® Industrial Biosciences, Brabrand Denmark) (SEQ ID NO:24). The latter is a neutral endo-protease. It is seen from Figure 4A that the combination of AteGlt2, FoodPro® PNL and AcPepN2 gives a large increase in the glutamic acid level in the final sausage. The increase is 0.65 mg / g of sausage relative to the blank (no enzymes added), corresponding to a 33-fold increase. The effect of AteGlt2 is seen by comparing to the trial where it has been omitted. A drop of 53% in glutamic acid level is observed relative to the trial where all three enzymes are present. It is also seen that omitting other of the three enzymes gives substantially lower levels of glutamic acid. An issue with adding endo-protease to these types of products is that the texture is affected. It is seen from Figure 4B that the sausages to which FoodPro PNL were added have a hardness that is only about half of the reference. For this setup the reference is much harder than a typical commercial sausage, while the hardness of the sausages to which were added FoodPro® PNL are in the range of commercial products. Hence, it is possible to produce a plant-based sausage with a commercially relevant level of glutamic and a desirable texture using the combination of the enzymes AteGlt2, FoodPro® PNL and AcPepN2. It was not possible to make comparable sausages with Amano enzyme since this product contains high levels of an endo-protease side activity. Hence, when Amano enzyme was used in the same dosage level as AteGlt2, hardness dropped to around 400 g, giving an overall texture that is not relevant for a plant-based sausage. Results from the experiments are shown in Figures 4A-4B. Although the foregoing invention has been described in some detail by way of illustration and example, for purposes of clarity of understanding, certain changes and NB41870-WO-PCT[3] modifications can be practiced within the scope of the appended claims. In addition, each reference provided herein is incorporated by reference in its entirety for all purposes to the same extent as if each reference was individually incorporated by reference. To the extent the content of any citation, including website or accession number may change with time, the version in effect at the filing date of this application is meant. Unless otherwise apparent from the context any step, element, aspect, feature of embodiment can be used in combination with any other.

[0002] NB41870-WO-PCT[3] SEQUENCE IDs: SEQ ID NO:1 AteGlt2 precursor protein; PRT; Aspergillus terreus: MLPLPLHWATALTALLCLGGASSANGDISAAGRFTTLSGIPYHVGGIAVSKIRDIS DVPSYPGALASIDVFPMTVISTSSFSYTGNELNSTIYDYLQKDDVFGIDFLHAIYLD SDGPGRCSIDSKSLRPELRQKAVALFMTSPRCSSGKPSIAVSKRLTHELPPGPYFV SAKTGEVFKAYRLYEDTHYAFLEPAVSDEQGGYLPLPSTTNSMLGPSIAVPSRLY YKPTAEQPLAGLRLGVKDIYHVKGLRTSGGNRAYYSLYEPRNATGPAVQRLVD QGAVFVGKMGTVQFANGDRPTADWVDFHCPFNPRGDGYLVPSGSSSGSGAGM GAYDWLDIAIGSDTGGSVRGPAGAQGLFGNRPSTGAVSMDNVIPLCDGLDTAGV FARDAHTWSTVVHAWYQDFDRNYTSFPKTLLYPSSSFSEGAIGSSDAAELIDNFV VNLERFLDTNRTDVDLDASWNTTRPSGTPATLNEMLHYTYGTLISVYQWLHHG QPFFEDYAAKHDGRTPYINPGPLLRWRLGQQSGQAGFDEAWNNKTNFYDWWN SDTGFGAHNAETCSESIYIYPNSVGGITYRDEYYSEPQAPYWGMSDSRIAVFAGV PDLVVPIGEVPYNSTKSGKTEYAPVTMSIVAARGCDLMVANMVAAMEEQGIVK PVAVGPRLYP SEQ ID NO:2 AteGlt2 predicted mature; PRT; Aspergillus terreus: ANGDISAAGRFTTLSGIPYHVGGIAVSKIRDISDVPSYPGALASIDVFPMTVISTSSF SYTGNELNSTIYDYLQKDDVFGIDFLHAIYLDSDGPGRCSIDSKSLRPELRQKAVA LFMTSPRCSSGKPSIAVSKRLTHELPPGPYFVSAKTGEVFKAYRLYEDTHYAFLEP AVSDEQGGYLPLPSTTNSMLGPSIAVPSRLYYKPTAEQPLAGLRLGVKDIYHVKG LRTSGGNRAYYSLYEPRNATGPAVQRLVDQGAVFVGKMGTVQFANGDRPTAD WVDFHCPFNPRGDGYLVPSGSSSGSGAGMGAYDWLDIAIGSDTGGSVRGPAGA QGLFGNRPSTGAVSMDNVIPLCDGLDTAGVFARDAHTWSTVVHAWYQDFDRN YTSFPKTLLYPSSSFSEGAIGSSDAAELIDNFVVNLERFLDTNRTDVDLDASWNTT RPSGTPATLNEMLHYTYGTLISVYQWLHHGQPFFEDYAAKHDGRTPYINPGPLL RWRLGQQSGQAGFDEAWNNKTNFYDWWNSDTGFGAHNAETCSESIYIYPNSVG GITYRDEYYSEPQAPYWGMSDSRIAVFAGVPDLVVPIGEVPYNSTKSGKTEYAPV TMSIVAARGCDLMVANMVAAMEEQGIVKPVAVGPRLYP SEQ ID NO:3 AteGlt2 native precursor DNA; DNA; Aspergillus terreus: ATGCTTCCTCTCCCTCTGCACTGGGCCACCGCCCTCACTGCCCTCCTGTGCCT GGGCGGCGCTAGCAGCGCTAACGGCGACATTAGCGCCGCTGGCCGATTCACC ACCCTGTCCGGCATCCCTTACCACGTTGGCGGCATCGCCGTTTCTAAGATCCG AGACATCTCCGACGTCCCCAGCTACCCCGGCGCTCTCGCTTCTATTGACGTTT TCCCCATGACCGTCATTAGCACTAGCTCTTTCAGCTACACCGGCAACGAGCTT AACAGCACTATTTACGACTACCTGCAGAAGGACGACGTTTTCGGCATTGACTT CCTGCACGCCATTTACCTTGACAGCGACGGCCCCGGCCGATGCAGCATTGAC AGCAAGTCTCTTCGACCTGAGCTCCGCCAGAAGGCTGTTGCTCTTTTCATGAC TAGCCCTCGATGCAGCAGCGGCAAGCCTTCTATCGCCGTGTCCAAGCGACTC ACCCACGAGCTGCCTCCCGGCCCTTACTTTGTCTCCGCTAAGACTGGCGAGGT TTTCAAGGCTTACCGCCTGTACGAGGACACTCACTACGCTTTCCTTGAGCCCG CTGTATCTGACGAGCAGGGCGGCTACCTGCCTCTCCCTAGCACTACTAACTCC ATGCTGGGCCCTTCTATTGCTGTTCCTAGCCGACTTTACTACAAGCCCACTGC TGAGCAGCCTCTTGCTGGCCTGCGACTTGGCGTTAAGGACATTTACCACGTCA NB41870-WO-PCT[3] AGGGCCTGCGAACTAGCGGCGGCAACCGCGCTTACTACAGCCTGTACGAGCC TCGAAACGCCACCGGCCCCGCTGTTCAGCGCCTTGTTGACCAGGGCGCCGTTT TCGTCGGCAAGATGGGCACTGTTCAGTTCGCTAACGGCGACCGACCTACTGC TGACTGGGTTGACTTCCACTGCCCTTTCAACCCCCGCGGCGACGGCTACCTTG TCCCCAGCGGCTCCAGCTCTGGCTCTGGCGCCGGCATGGGCGCCTACGACTG GCTTGACATTGCCATTGGCTCCGACACTGGCGGCAGCGTCCGAGGCCCCGCC GGCGCTCAGGGCCTTTTCGGCAACCGACCTAGCACTGGCGCTGTTAGCATGG ACAACGTTATTCCTCTGTGCGACGGCCTGGACACCGCTGGCGTTTTCGCACGT GACGCTCACACTTGGTCCACCGTCGTTCACGCTTGGTATCAGGACTTCGACCG AAACTACACTTCTTTCCCCAAGACTCTTCTGTACCCTAGCTCTAGCTTCTCTGA GGGCGCCATCGGCAGCTCCGACGCTGCTGAGCTTATTGACAACTTCGTTGTCA ACCTTGAGCGATTCCTGGACACCAACCGAACTGACGTCGACCTGGACGCTTC TTGGAACACCACTCGACCTAGCGGCACTCCCGCCACCCTTAACGAGATGCTG CACTACACTTACGGCACCCTGATTAGCGTTTACCAGTGGCTCCACCACGGCCA GCCTTTCTTCGAGGACTACGCTGCTAAGCACGACGGCCGAACTCCTTACATTA ACCCCGGCCCTCTTCTGCGATGGCGCCTGGGCCAGCAGTCTGGCCAGGCTGG CTTCGACGAGGCTTGGAACAACAAGACCAACTTCTACGACTGGTGGAACTCT GACACTGGCTTCGGCGCTCACAACGCTGAGACTTGCTCTGAGTCTATCTACAT TTACCCCAACAGCGTCGGCGGCATCACCTACCGAGACGAGTACTACTCTGAG CCTCAGGCTCCTTACTGGGGCATGTCTGACTCTCGAATTGCCGTTTTCGCCGG CGTTCCTGACCTTGTTGTTCCTATTGGCGAGGTTCCCTACAACTCCACCAAGT CCGGCAAGACTGAGTACGCCCCCGTCACCATGTCCATCGTTGCTGCGAGGGG CTGCGACCTTATGGTCGCTAACATGGTTGCCGCTATGGAGGAGCAGGGCATC GTTAAGCCCGTTGCCGTTGGCCCTCGCCTGTACCCC SEQ ID NO:4 EcoGlt1 full length; PRT; Escherichia coli: MSDAKNLQQAVDQAYVQFHSLSGGQNADYIPFLANVPSQLAGVAIVTCDGNVY RAGDSDYRFALESISKVCTLALALEDVGPQAVQDKIGADPTGLPFNSVIALELHG GKPLSPLVNAGAIATTSLIKADNAEQRWQRILHIQQQLAGEQVALSEEVNQSEQT TNFHNRAIAWLLYSSGYLYCDAMEACDVYTRQCSTLINTVELATLGATLAAGGV NPLSQKRVLQADNVPYILAEMMMEGLYGRSGDWAYRVGLPGKSGVGGGILAV VPGVMGIAAFSPPLDEEGNSVRGQKMVASVAKQLGYNVFKG SEQ ID NO:5 EcoGlt1 synthetic / optimized full length DNA; DNA; Synthetic: ATGAGCGATGCAAAAAATCTTCAACAAGCAGTTGATCAAGCATATGTTCAAT TTCATTCTCTGAGCGGAGGCCAAAATGCAGATTATATCCCTTTTCTTGCAAAT GTTCCGAGCCAACTGGCAGGAGTTGCAATTGTTACGTGCGATGGAAATGTTT ATCGCGCAGGAGATTCTGATTATCGCTTTGCACTTGAATCTATTTCTAAAGTT TGCACGCTGGCACTGGCACTTGAAGATGTTGGACCTCAAGCAGTTCAAGATA AAATCGGCGCAGATCCTACGGGCTTACCTTTTAATAGCGTTATCGCATTAGAA TTACATGGCGGAAAACCGCTGTCACCTCTGGTTAATGCAGGCGCAATCGCAA CGACATCCCTTATTAAAGCAGATAATGCAGAACAACGCTGGCAAAGAATCCT TCATATCCAACAACAACTGGCAGGAGAACAAGTTGCACTGTCTGAAGAAGTT AATCAAAGCGAACAAACGACGAATTTTCATAATAGAGCAATCGCATGGTTAC TGTATAGCTCAGGCTATCTGTATTGCGATGCAATGGAAGCATGCGATGTTTAT ACGAGACAATGCTCTACACTGATCAATACGGTTGAACTTGCAACACTGGGCG NB41870-WO-PCT[3] CAACGTTAGCAGCAGGAGGCGTTAATCCGCTGTCACAAAAACGCGTTTTACA AGCAGATAATGTTCCGTATATCTTAGCAGAAATGATGATGGAAGGCCTGTAT GGACGCTCAGGAGATTGGGCATATAGAGTTGGACTTCCGGGAAAATCAGGCG TTGGCGGCGGTATACTGGCAGTTGTTCCGGGAGTTATGGGCATCGCAGCATTT AGCCCGCCTCTTGATGAAGAAGGAAATAGCGTTCGCGGCCAAAAAATGGTTG CATCAGTTGCAAAACAATTAGGCTATAATGTTTTTAAAGGA SEQ ID NO:6 MetGlt2 precursor; PRT; Unidentified: MKRTWNVCLTALLSVLLVAGSVPFHAEAKKPPKSYDEYKQVDVGKDGMVATA HPLASEIGADVLKKGGNAIDAAVAIQFALNVTEPMMSGIGGGGFMMVYDGKTK DTTIIDSRERAPAGATPDMFLDENGKAIPFSERVTKGTAVGVPGTLKGLEEALDK WGTRSMKQLITPSIKLAEKGFPIDSVLAEAISDYQEKLSRTAAKDVFLPNGEPLKE GDTLIQKDLAKTFKLIRSKGTDAFYKGKFAKALSDTVQDFGGSMTEKDLENYDI TIDEPIWGDYQGYQIATTPPPSSGGIFLLQMLKILDHFNLSQYDVRSWEKYQLLAE TMHLSYADRASYAGDPEFVNVPLKGLLHPDYIKERQQLINLDQVNKKPKAGDP WKYQEGSANYKQVEQPKDKVEGQTTHFTVADRWGNVVSYTTTIEQLFGTGIMV PDYGVILNNELTDFDAIPGGANEVQPNKRPLSSMTPTILFKDDKPVLTVGSPGGA TIISSVLQTILYHIEYGMELKAAVEEPRIYTNSMSSYRYEDGVPKDVLSKLNGMG HKFGTSPVDIGNVQSISIDHENGTFKGVADSSRNGAAIGINLKRK SEQ ID NO:7 MetGlt2 predicted mature; PRT; Unidentified: KKPPKSYDEYKQVDVGKDGMVATAHPLASEIGADVLKKGGNAIDAAVAIQFAL NVTEPMMSGIGGGGFMMVYDGKTKDTTIIDSRERAPAGATPDMFLDENGKAIPF SERVTKGTAVGVPGTLKGLEEALDKWGTRSMKQLITPSIKLAEKGFPIDSVLAEA ISDYQEKLSRTAAKDVFLPNGEPLKEGDTLIQKDLAKTFKLIRSKGTDAFYKGKF AKALSDTVQDFGGSMTEKDLENYDITIDEPIWGDYQGYQIATTPPPSSGGIFLLQ MLKILDHFNLSQYDVRSWEKYQLLAETMHLSYADRASYAGDPEFVNVPLKGLL HPDYIKERQQLINLDQVNKKPKAGDPWKYQEGSANYKQVEQPKDKVEGQTTHF TVADRWGNVVSYTTTIEQLFGTGIMVPDYGVILNNELTDFDAIPGGANEVQPNK RPLSSMTPTILFKDDKPVLTVGSPGGATIISSVLQTILYHIEYGMELKAAVEEPRIY TNSMSSYRYEDGVPKDVLSKLNGMGHKFGTSPVDIGNVQSISIDHENGTFKGVA DSSRNGAAIGINLKRK SEQ ID NO:8 MetGlt2 optimized / synthetic precursor DNA; DNA; Synthetic: GTGAGAAGCAAAAAATTGTGGATCAGCTTGTTGTTTGCGTTAACGTTAATCTT TACGATGGCGTTCAGCAACATGAGCGCGCAGGCTAAAAAACCGCCGAAATC ATATGATGAATATAAACAAGTTGATGTTGGCAAAGATGGCATGGTTGCAACA GCACATCCGTTAGCATCAGAAATCGGCGCAGATGTTCTTAAAAAAGGAGGAA ATGCAATTGATGCAGCAGTTGCAATCCAATTTGCACTTAATGTTACAGAACCT ATGATGTCAGGAATCGGCGGCGGAGGCTTTATGATGGTTTATGATGGCAAAA CGAAAGATACGACAATCATCGATAGCCGCGAACGCGCACCTGCAGGCGCAA CACCGGATATGTTTCTGGATGAAAATGGCAAAGCAATCCCTTTTAGCGAACG CGTTACGAAAGGAACAGCAGTTGGAGTTCCGGGCACACTGAAAGGCCTGGA AGAAGCACTTGATAAATGGGGCACGAGAAGCATGAAACAACTTATTACGCCG TCTATTAAACTTGCAGAAAAAGGCTTTCCGATCGATAGCGTTTTAGCAGAAG CAATTAGCGATTATCAAGAAAAACTGTCTCGCACAGCAGCAAAAGATGTTTT NB41870-WO-PCT[3] TCTGCCTAATGGAGAACCGCTTAAAGAAGGAGATACACTTATCCAAAAAGAT CTTGCAAAAACGTTTAAACTTATTCGCTCTAAAGGCACGGATGCATTTTATAA AGGCAAATTTGCAAAAGCACTGAGCGATACAGTTCAAGATTTTGGCGGCTCT ATGACGGAAAAAGATTTAGAAAATTATGATATTACAATTGATGAACCGATCT GGGGCGATTATCAAGGCTATCAAATCGCAACGACGCCTCCTCCGAGCTCAGG AGGCATCTTTCTGCTTCAAATGCTGAAAATCCTGGATCATTTTAATCTGAGCC AATATGATGTTCGCTCTTGGGAAAAATATCAACTGCTTGCAGAAACAATGCA TCTGTCATATGCAGATAGAGCAAGCTATGCAGGAGATCCTGAATTTGTTAAT GTTCCTCTTAAAGGACTGCTTCATCCGGATTATATCAAAGAACGCCAACAACT TATTAATCTGGATCAAGTTAATAAAAAACCTAAAGCAGGAGATCCGTGGAAA TATCAAGAAGGCTCAGCAAATTATAAACAAGTTGAACAACCTAAAGATAAAG TTGAAGGCCAAACGACACATTTTACGGTTGCAGATCGCTGGGGAAATGTTGT TAGCTATACGACGACGATCGAACAACTGTTTGGCACGGGAATCATGGTTCCG GATTATGGAGTTATTCTTAATAATGAACTGACGGATTTTGATGCAATTCCGGG CGGCGCAAATGAAGTTCAACCTAATAAACGCCCGCTGAGCTCTATGACACCT ACGATTCTGTTTAAAGATGATAAACCGGTTCTGACGGTTGGCTCACCGGGCG GCGCAACGATCATCTCTAGCGTTCTTCAAACGATCCTGTATCATATCGAATAT GGCATGGAACTGAAAGCAGCAGTTGAAGAACCTCGCATCTATACGAATAGCA TGTCTAGCTATCGCTATGAAGATGGAGTTCCTAAAGATGTTCTGTCTAAACTG AATGGCATGGGACATAAATTTGGCACGTCACCGGTTGATATTGGAAATGTTC AATCTATCTCTATTGATCATGAAAATGGCACGTTTAAAGGCGTTGCAGATAGC TCACGCAATGGCGCAGCAATCGGCATTAATCTTAAACGCAAA SEQ ID NO:9 MetGlt7 precursor; PRT; Unidentified: MKRISLTVLSICLFVFSFFLPVSQVTANETHGNKVAVGKDGMVATAHPLASEIGA DVLKKGGNAVDAAVAIQYALNVTEPMMSGIGGGGFMMVYDGKTKETSIINSRE RAPQGATPDMFLTDDGKVIPFAERSTHGNAVGVPGTVKGLEAALDKWGTRSMK ELIEPSIQLAEDGFEIDSVLANAIDDHQGKLKKTAAAPIFLPNDQPLEEGDLLVQP GLAKTFKLIAKKGSKAFYEGKVAKALANTVQDFGGTMTSKDIKRYEVKTDKPI WGDYKGYQLASMPPPSSGGVFMLQILKILDHFNLSQYDPKSFEKYQLLAETMHL SYADRAAYAGDPEFVDVPLKGLLDDDYISERASLIQLDQMNRSPKEGDPWAYED EKNPSPIVPQPEDKTIGETTHFTVADQWGNVVSFTTTIEQLFGTGILVPEYGFFLN NELTDFDARPGGANEVQPNKRPLSSMTPTIIFKDGEPVMTVGSPGGTTIIASVSQTI LNLLEYDMELQDAVEEPRIYTNSLTSYRYEVGVPLDVRTKLNDMGHQFGSSPIDI GNVQALLIDRKAGTFTGVADSTRNGTAVGVNLKVAADQ SEQ ID NO:10 MetGlt7 predicted mature; PRT; Unidentified: NETHGNKVAVGKDGMVATAHPLASEIGADVLKKGGNAVDAAVAIQYALNVTE PMMSGIGGGGFMMVYDGKTKETSIINSRERAPQGATPDMFLTDDGKVIPFAERS THGNAVGVPGTVKGLEAALDKWGTRSMKELIEPSIQLAEDGFEIDSVLANAIDD HQGKLKKTAAAPIFLPNDQPLEEGDLLVQPGLAKTFKLIAKKGSKAFYEGKVAK ALANTVQDFGGTMTSKDIKRYEVKTDKPIWGDYKGYQLASMPPPSSGGVFMLQI LKILDHFNLSQYDPKSFEKYQLLAETMHLSYADRAAYAGDPEFVDVPLKGLLDD DYISERASLIQLDQMNRSPKEGDPWAYEDEKNPSPIVPQPEDKTIGETTHFTVAD QWGNVVSFTTTIEQLFGTGILVPEYGFFLNNELTDFDARPGGANEVQPNKRPLSS MTPTIIFKDGEPVMTVGSPGGTTIIASVSQTILNLLEYDMELQDAVEEPRIYTNSLT NB41870-WO-PCT[3] SYRYEVGVPLDVRTKLNDMGHQFGSSPIDIGNVQALLIDRKAGTFTGVADSTRN GTAVGVNLKVAADQ SEQ ID NO:11 MetGlt7 optimized / synthetic precursor DNA; DNA; Synthetic: GTGAGAAGCAAAAAATTGTGGATCAGCTTGTTGTTTGCGTTAACGTTAATCTT TACGATGGCGTTCAGCAACATGAGCGCGCAGGCTAATGAAACACATGGAAAT AAAGTTGCAGTTGGCAAAGATGGCATGGTTGCAACAGCACATCCGTTAGCAA GCGAAATCGGCGCAGATGTTCTGAAAAAAGGAGGAAATGCAGTTGATGCAG CAGTTGCAATCCAATATGCACTTAATGTTACAGAACCGATGATGTCAGGAAT CGGCGGCGGAGGCTTTATGATGGTTTATGATGGCAAAACGAAAGAAACGTCT ATTATTAATTCTCGCGAACGCGCACCTCAAGGCGCAACACCGGATATGTTTCT TACGGATGATGGCAAAGTTATCCCGTTTGCAGAACGCTCTACACATGGCAAT GCAGTTGGAGTTCCGGGCACGGTTAAAGGCCTGGAAGCAGCACTGGATAAAT GGGGAACGCGCTCTATGAAAGAACTTATCGAACCGTCTATCCAACTTGCAGA AGATGGCTTTGAAATTGATAGCGTTTTAGCAAATGCAATCGATGATCATCAA GGCAAACTTAAAAAAACAGCAGCAGCACCGATCTTTCTGCCTAATGATCAAC CGTTAGAAGAAGGAGATTTACTGGTTCAACCGGGCCTTGCAAAAACATTTAA ACTTATCGCAAAAAAAGGCTCTAAAGCATTTTATGAAGGCAAAGTTGCAAAA GCATTAGCAAATACGGTTCAAGATTTTGGAGGAACAATGACGTCTAAAGATA TTAAACGCTATGAAGTTAAAACGGATAAACCGATCTGGGGCGATTATAAAGG CTATCAACTTGCATCTATGCCGCCGCCGAGCTCAGGAGGCGTTTTTATGCTTC AAATTCTTAAAATCCTGGATCATTTTAATCTGAGCCAATATGATCCTAAATCA TTTGAAAAATATCAACTGCTGGCAGAAACAATGCATCTGTCATATGCAGATA GAGCAGCATATGCAGGCGATCCGGAATTTGTTGATGTTCCTCTGAAAGGACT GCTGGATGATGATTATATTAGCGAACGCGCATCACTTATCCAACTGGATCAA ATGAATCGCTCACCTAAAGAAGGAGATCCGTGGGCATATGAAGATGAAAAA AATCCGAGCCCTATTGTTCCTCAACCTGAAGATAAAACAATCGGCGAAACGA CACATTTTACGGTTGCAGATCAATGGGGAAATGTTGTTAGCTTTACGACGACG ATCGAACAACTGTTTGGCACAGGGATACTTGTTCCGGAATATGGCTTTTTTCT GAATAATGAACTGACGGATTTTGATGCACGACCTGGCGGCGCAAATGAAGTT CAACCTAATAAACGCCCGCTGTCTAGCATGACACCTACGATCATCTTTAAAG ATGGAGAACCTGTTATGACGGTTGGCTCACCGGGAGGCACGACAATTATCGC AAGCGTTTCACAAACGATTCTTAATCTGCTTGAATATGATATGGAATTACAAG ATGCAGTTGAAGAACCTCGCATCTATACGAATAGCCTGACGAGCTATCGCTA TGAAGTTGGAGTTCCGCTGGATGTTCGCACGAAACTTAATGATATGGGACAT CAATTTGGCTCTAGCCCTATTGATATTGGAAATGTTCAAGCACTGCTTATCGA TAGAAAAGCAGGCACGTTTACAGGCGTTGCAGATTCTACACGCAATGGCACA GCAGTTGGCGTTAATCTTAAAGTTGCAGCAGATCAA SEQ ID NO:12 MspGlt2 precursor; PRT; Melanconium sp. NRRL 54901: MRSALLSLVIIGHASGLAVDKGRVIDLQGVSYYAGGTPIARLDINGTREAITAAAP DGHDLFPLTVVETSSGSFSAQKIVNITAEYDEMDDVFQPAFLQTVYLQPVDDNNI FAIDITSAKSVLDELEMKLLIARGNASTQSPSVVTGTITPSLPKGPYFVSAYSGDV YKAYRLYDDNNLAFIQGIISDDQGAYMSLPAVIENVMAKSIAVPSRLYYNATKE KPLAGLRFGVKDIFHVKGITTSGGNRAYFYHYGTQNNTAPAVQRLIDLGAVLIGK NB41870-WO-PCT[3] MGTVQFANGDRPTADWVDLHAPFNPRGDGYQDPSGSSTGPGAGIGAYEWLDLS VGSDTGGSMRGPAGAQGIFGNRPSTGAISLDHVIPLSPVSDTAGLFARSGRLWAR ATKAYYPSLQSNWTAFPKKLYSSPVTNGTSADAVALVNTFVQKLGDLLGTEALP GNYTQLWAETAGNAPAVDEMLNLTYAVFISHDQWRMLGKPFFEDYAAKNGGR QPYINPGPLARWQWGQINAPDEVYAEALRNISTFTEWYTTEGYGRHDPESCSESV YTYLFRSGQPSYRDEYFSAPTSPPFGMSDSRVAVIAGAPEVIVPIGEVPYNSTKSL QVEYLPVSMALRMARGCDYVLADLVDKLEEQGIVRPVAAGSRLYP SEQ ID NO:13 MspGlt2 predicted mature; PRT; Melanconium sp. NRRL 54901 LAVDKGRVIDLQGVSYYAGGTPIARLDINGTREAITAAAPDGHDLFPLTVVETSS GSFSAQKIVNITAEYDEMDDVFQPAFLQTVYLQPVDDNNIFAIDITSAKSVLDELE MKLLIARGNASTQSPSVVTGTITPSLPKGPYFVSAYSGDVYKAYRLYDDNNLAFI QGIISDDQGAYMSLPAVIENVMAKSIAVPSRLYYNATKEKPLAGLRFGVKDIFHV KGITTSGGNRAYFYHYGTQNNTAPAVQRLIDLGAVLIGKMGTVQFANGDRPTAD WVDLHAPFNPRGDGYQDPSGSSTGPGAGIGAYEWLDLSVGSDTGGSMRGPAGA QGIFGNRPSTGAISLDHVIPLSPVSDTAGLFARSGRLWARATKAYYPSLQSNWTA FPKKLYSSPVTNGTSADAVALVNTFVQKLGDLLGTEALPGNYTQLWAETAGNA PAVDEMLNLTYAVFISHDQWRMLGKPFFEDYAAKNGGRQPYINPGPLARWQW GQINAPDEVYAEALRNISTFTEWYTTEGYGRHDPESCSESVYTYLFRSGQPSYRD EYFSAPTSPPFGMSDSRVAVIAGAPEVIVPIGEVPYNSTKSLQVEYLPVSMALRM ARGCDYVLADLVDKLEEQGIVRPVAAGSRLYP SEQ ID NO:14 MspGlt2 native precursor DNA; DNA; Melanconium sp. NRRL 54901: ATGCGGTCTGCCCTGCTCAGTCTGGTTATCATCGGCCATGCGTCCGGGCTGGC CGTTGACAAGGGGCGTGTCATCGATTTACAAGGAGTGTCATACTATGCTGGT GGCACTCCGATCGCCCGTCTCGATATCAATGGCACTCGTGAAGCGATCACCG CCGCCGCCCCCGATGGTCACGATTTGTTTCCGCTGACCGTTGTCGAGACCTCA TCAGGGAGCTTCAGTGCCCAGAAAATCGTTAATATCACCGCTGAGTACGATG AGATGGACGACGTCTTCCAGCCGGCCTTCCTCCAGACTGTCTACCTGCAGCCC GTAGACGACAATAATATTTTTGCAATTGATATCACGTCTGCGAAATCCGTTCT GGATGAGTTGGAGATGAAGCTCCTAATCGCCCGCGGCAACGCATCGACACAA TCTCCATCTGTCGTGACCGGCACGATCACTCCTTCTCTCCCAAAGGGTCCTTA TTTCGTCTCAGCCTACAGTGGTGATGTCTATAAGGCTTACCGGCTGTACGATG ACAACAACCTTGCTTTCATCCAGGGAATTATCAGCGACGATCAGGGTGCCTA CATGTCCCTGCCAGCTGTCATAGAGAACGTCATGGCTAAGAGTATCGCTGTG CCATCCAGGCTCTACTACAATGCCACCAAGGAGAAACCCCTTGCCGGTCTGC GTTTCGGCGTCAAGGATATTTTCCATGTCAAGGGAATCACTACCAGCGGCGG CAATCGTGCCTATTTCTATCACTACGGTACTCAAAACAACACGGCTCCGGCCG TGCAGCGCCTCATCGACCTCGGTGCCGTCCTCATCGGCAAGATGGGAACTGT TCAATTTGCCAACGGCGATAGGCCTACTGCTGACTGGGTTGACCTGCATGCCC CATTCAACCCTCGGGGTGACGGCTACCAAGACCCCAGCGGCTCTTCCACTGG GCCTGGTGCCGGAATAGGTGCTTATGAATGGCTGGATCTGTCCGTTGGTAGTG ATACTGGCGGTTCAATGCGCGGTCCTGCAGGAGCCCAAGGCATCTTCGGCAA CAGACCCTCGACCGGTGCCATCTCGCTAGACCATGTCATCCCTCTCAGCCCTG TGTCCGACACAGCCGGCCTGTTCGCCCGCAGCGGAAGGCTCTGGGCCAGGGC GACGAAAGCTTACTATCCCAGCTTGCAATCCAACTGGACCGCGTTTCCCAAG NB41870-WO-PCT[3] AAGCTCTACAGCTCGCCTGTCACCAACGGTACAAGCGCCGACGCTGTAGCGC TGGTAAATACCTTCGTCCAGAAGCTGGGCGACTTGCTCGGAACAGAGGCGCT CCCGGGCAACTATACCCAGCTGTGGGCCGAAACTGCCGGCAATGCGCCCGCC GTCGACGAGATGCTGAACCTGACATACGCGGTCTTCATATCCCACGACCAGT GGCGGATGCTCGGCAAGCCCTTCTTCGAGGACTACGCCGCTAAGAATGGCGG CCGCCAGCCGTATATCAACCCGGGGCCCCTGGCACGCTGGCAGTGGGGACAG ATCAACGCCCCTGATGAGGTCTATGCAGAGGCACTCCGTAACATTTCCACCTT CACAGAGTGGTACACCACCGAGGGCTACGGCCGCCACGACCCAGAGTCCTGC TCCGAGAGTGTGTACACCTACCTCTTCCGGAGCGGCCAGCCCTCGTACCGTG ATGAATACTTCTCGGCCCCTACGAGCCCTCCTTTTGGCATGAGCGATTCACGG GTTGCTGTTATTGCTGGGGCTCCTGAGGTCATCGTCCCTATTGGCGAGGTGCC ATATAACAGCACAAAGTCCCTGCAGGTAGAGTATTTACCTGTGTCAATGGCA CTCAGAATGGCGAGGGGCTGCGACTACGTCCTTGCCGACTTAGTAGACAAGC TGGAGGAGCAGGGAATTGTTCGACCTGTTGCTGCTGGCTCTAGATTATATCCT TAA SEQ ID NO:15 EtaGlt1 full length; PRT; Edwardsiella tarda: MTLEAAALQQAVDQAHRQYGALPGGKNADYIPYLAGVPSQLAAVAIVTRDGAV YCAGDSAYRFALESISKVCTLALALEDVGPQAVQDKIGADPTGLPFNSVMALEL HGGKPLSPLVNAGAMAAASLIQAPDREQRWQRILAIQQQLAGEKVALSDEVNQS EQTTNFHNRAIAWLLYSAGTMYCDPMEACDVYTRQCSTLIDTVELATLGATLAA GGVNPRSGQRVLQADNVPYILAEMTMEGLYGRSGDWAYHVGLPGKSGVGGGIL AVVPGVMGIAAFSPPLDEAGNSVRGQKMVAAVAAQLGYNLFKA SEQ ID NO:16 EtaGlt1 synthetic / optimized full length DNA; DNA; Synthetic: GTGACACTGGAAGCAGCAGCACTGCAACAAGCAGTTGATCAAGCACATAGA CAATATGGCGCACTGCCGGGCGGCAAAAATGCAGATTATATTCCGTATCTGG CAGGCGTTCCGTCACAACTGGCAGCAGTTGCGATTGTTACAAGAGATGGCGC AGTTTATTGCGCAGGCGATAGCGCGTATAGATTTGCACTGGAATCAATTTCAA AAGTTTGCACACTTGCACTGGCACTTGAAGATGTTGGCCCGCAAGCAGTTCA AGATAAAATTGGCGCAGATCCGACAGGCCTGCCGTTTAATAGCGTTATGGCA CTGGAACTTCATGGCGGCAAACCGTTATCACCGCTTGTTAATGCAGGCGCGA TGGCGGCAGCATCACTTATTCAAGCACCGGATAGAGAACAAAGATGGCAAA GAATTCTGGCAATTCAACAACAACTGGCAGGCGAAAAAGTTGCACTGTCAGA TGAAGTTAATCAATCAGAACAAACAACAAATTTTCATAATAGAGCAATTGCA TGGCTGCTGTATTCAGCAGGCACAATGTATTGCGATCCGATGGAAGCATGCG ATGTTTATACAAGACAATGCAGCACACTGATTGATACAGTTGAACTGGCAAC ACTGGGCGCGACACTGGCAGCAGGCGGCGTTAATCCGAGATCAGGCCAAAG AGTTCTTCAAGCAGATAATGTTCCGTATATTCTGGCAGAAATGACAATGGAA GGCCTGTATGGCAGATCAGGCGATTGGGCATATCATGTTGGCCTGCCGGGCA AAAGCGGCGTTGGCGGCGGAATTCTTGCGGTTGTTCCGGGCGTTATGGGCAT TGCGGCATTTTCACCGCCGCTGGATGAAGCAGGCAATAGCGTTAGAGGCCAA AAAATGGTTGCAGCAGTTGCAGCACAACTTGGCTATAATCTTTTTAAAGCATA A SEQ ID NO:17 TspGlt1 full length; PRT; Tatumella sp.: NB41870-WO-PCT[3] MQQDPQVIRQSIEQALQESLSLQGGKNADYIPFLAQVPSHLSAVAVVTAEGAVY QAGDSSYPFAIESISKVSTLALALEEAGPAAIKEKIGADPTGLPFNSVIALELHSGK PLSPLVNAGAMSAVSLLKANDAEQRWQRILHLQQQMAGSALALSEEVNQSEQS TNFHNRGIAWLLYSAGYMYCDPMEACEVYTRQCSVLINTTQLATFAATLAARG MNPLTRQQVLRKDNVPCILAEMTMEGMYGASGDWAYSVGLPAKSGVGGGVIA VVPGVMGIAAFSPPLDNEGNSVRAQAMVASVASQLGYSLYR SEQ ID NO:18 TspGlt1 synthetic / optimized full length DNA; DNA; Synthetic: GTGCAACAAGATCCGCAAGTTATTAGACAATCAATTGAACAAGCGCTGCAAG AAAGCCTGAGCCTGCAAGGCGGCAAAAATGCAGATTATATTCCGTTTCTGGC ACAAGTTCCGTCACATCTGAGCGCAGTTGCAGTTGTTACAGCGGAAGGCGCA GTTTATCAAGCAGGCGATTCATCATATCCGTTTGCAATTGAATCAATTAGCAA AGTTAGCACACTGGCACTTGCACTGGAAGAAGCAGGCCCGGCAGCGATTAAA GAAAAAATTGGCGCAGATCCGACAGGCCTGCCGTTTAATTCAGTTATTGCAC TTGAACTGCATAGCGGCAAACCGCTGAGCCCGCTGGTTAATGCAGGCGCAAT GAGCGCAGTTAGCCTGCTTAAAGCAAATGATGCGGAACAAAGATGGCAAAG AATTCTTCATCTGCAACAACAAATGGCAGGCTCAGCACTGGCACTTAGCGAA GAAGTTAATCAAAGCGAACAATCAACAAATTTTCATAATAGAGGCATTGCAT GGCTTCTGTATAGCGCAGGCTATATGTATTGCGATCCGATGGAAGCATGCGA AGTGTATACAAGACAATGCTCTGTTCTGATTAATACAACACAACTGGCAACA TTTGCAGCGACACTGGCAGCAAGAGGCATGAATCCGCTTACAAGACAACAAG TTCTGAGAAAAGATAATGTTCCGTGCATTCTGGCAGAAATGACAATGGAAGG CATGTATGGAGCAAGCGGAGATTGGGCATATTCAGTTGGCCTGCCGGCGAAA TCAGGCGTTGGCGGCGGCGTTATTGCAGTTGTTCCGGGCGTTATGGGCATTGC GGCATTTTCACCGCCGCTGGATAATGAAGGCAATAGCGTTAGAGCACAAGCA ATGGTTGCATCAGTTGCGAGCCAACTGGGCTATAGCCTGTATAGATAA SEQ ID NO:19 SenGlt2 full length; PRT; Salmonella enterica: MSDNHELIQRAVTNAWQQFSTLEGGEKASYIPFLANVPGNLSAVAVVTTRGEIFT EGEASYRFALESISKVYTLALALEDVGPSEVQNKIGADPTGLPFNSVLALELHHG RPLSPLVNAGAMAAVSLIKASDREERWARILDIQRQLAGAPVMLSDEINRSEQDT NFHNRAIAWLLYSAGTLYCDPMEACDVYTRQCSTLMNTTELATTGATLAAGGI NPCNGSRVLSPSSIPYILAEMTMEGMYGSSGDWAYTVGLPGKSGVGGGILAVVP GVMGIAAFSPPLDGSGNSVRGQKMVASVAKELGYNLYDIR SEQ ID NO:20 SenGlt2 synthetic / optimized full length DNA; DNA; Synthetic: GTGAGCGATAATCATGAACTGATTCAAAGAGCAGTTACAAATGCATGGCAAC AATTTTCAACACTTGAAGGCGGCGAAAAAGCAAGCTATATTCCGTTTCTGGC AAATGTTCCGGGCAATCTGAGCGCGGTTGCGGTTGTGACAACAAGAGGCGAA ATTTTTACAGAAGGCGAAGCATCATATAGATTTGCGCTGGAAAGCATTAGCA AAGTTTATACACTTGCGCTTGCACTGGAAGATGTTGGCCCGTCAGAAGTGCA AAATAAAATTGGCGCAGATCCGACAGGCCTTCCGTTTAATAGCGTTCTGGCA CTGGAACTGCATCATGGCAGACCGCTTAGCCCGCTGGTTAATGCAGGCGCGA TGGCAGCAGTGTCACTGATTAAAGCGTCAGATAGAGAAGAAAGATGGGCGA GAATTCTGGATATTCAAAGACAACTTGCAGGCGCACCGGTTATGCTGTCTGAT GAAATTAATAGATCAGAACAAGATACAAATTTTCATAATAGAGCAATTGCGT NB41870-WO-PCT[3] GGCTTCTGTATAGCGCAGGCACACTGTATTGCGATCCGATGGAAGCATGCGA TGTTTATACAAGACAATGCTCAACACTGATGAATACAACAGAACTGGCAACA ACAGGCGCAACACTGGCAGCAGGCGGCATTAATCCGTGCAATGGCAGCAGA GTTCTGAGCCCGAGCAGCATTCCGTATATTCTGGCAGAAATGACAATGGAAG GCATGTATGGATCATCAGGCGATTGGGCGTATACAGTTGGCCTTCCGGGCAA ATCAGGCGTTGGCGGCGGCATTCTGGCAGTTGTTCCGGGCGTTATGGGCATTG CAGCATTTAGCCCGCCGCTGGATGGCAGCGGCAATTCAGTTAGAGGACAAAA AATGGTTGCAAGCGTTGCAAAAGAACTGGGCTATAATCTGTATGATATTAGA TAA SEQ ID NO:21 Amano_SD-C100S_glutaminase precursor; PRT; Bacillus amyloliquefaciens: MKKKKFMNLCFIVLLSALLTAGSIPYHAQAKKHPFSYDDYKQVDVGKDGMVAT AHPLASQIGADVLKKGGNAIDAAVAIQFALNVTEPMMSGIGGGGFMMVYDAKT KDTTIIDSRERAPAGATPDMFLDENGKAIPFSERVTKGTAVGVPGTLKGLEKALD KWGTRSMKQLITPSIALASKGFPIDSVLADAISDYKDKLSHTAAKDVFLPNGEPL KEGDTLVQKDLAKTFTAIKYKGTKAFYDGAFTKKLAETVQEFGGSMTEQDIKNF NVTIDEPIWGDYQGYHIATAPPPSSGGVFLLQMLNLLDDFKLSQYDIRSWQKYQL LAETMHLAYADRAAFAGDPEFVNVPLKGLLNPDYINARRQLIDINKVNKKPKAG DPWAYQEGSANYKQVEQPTDKQEGQTTHFTVADRFGNVVSYTTTIEQLFGSGIM VPGYGVVLNNELTDFDAVPGGANEVQPNKRPLSSMTPTILFKNNEPVLTVGSPG GATIISSVLQTILNKVEYGMDLKAAVEEPRIYTNSMTSYRYEEGVPEEARTKLNE MGHKFGSKPVDIGNVQSILIDRENGTFTGVADSSRNGAAIGVNLKKCEK SEQ ID NO:22 subtilisin like alkaline protease precursor; PRT; Bacillus licheniformis: MMRKKSFWLGMLTAFMLVFTMAFSDSASAAQPAKNVEKDYIVGFKSGVKTAS VKKDIIKESGGKVDKQFRIINAAKAKLDKEALKEVKNDPDVAYVEEDHVAHALA QTVPYGIPLIKADKVQAQGFKGANVKVAVLDTGIQASHPDLNVVGGASFVAGEA YNTDGNGHGTHVAGTVAALDNTTGVLGVAPSVSLYAVKVLNSSGSGSYSGIVS GIEWATTNGMDVINMSLGGASGSTAMKQAVDNAYARGVVVVAAAGNSGSSGN TNTIGYPAKYDSVIAVGAVDSNSNRASFSSVGAELEVMAPGAGVYSTYPTNTYA TLNGTSMASPHVAGAAALILSKHPNLSASQVRNRLSSTATYLGSSFYYGKGLINV EAAAQ SEQ ID NO:23 AcPepN2 N-terminal exopeptidase precurosr; PRT; Aspergillus clavatus: MKWLYLAAFASLALANAPGGPGGHGRKLPVNPKTFPNEIRLKDLLHGSQKLEDF AYAYPERNRVFGGQAHLDTVNYLYRELKKTGYYDVYKQPQVHQWTRADQSLT LGGDSIQASTMTYSPSVNVTAPLSLVSKLGCAEGDYSADVKGKIALVSRGECSFA QKSVLSAKAGAVATIVYNNVDGSLAGTLGGATSELGPYSPIIGITLAAGQDLVAR LQAAPTEVSLWIDSKVENRTTYNVIAQTKGGDPNNVVALGGHTDSVENGPGIND DGSGVISNLVVAKALTRYSVKNAVRFCFWTAEEFGLLGSNYYVDNLSPAELAKI RLYLNFDMIASPNYALMIYDGDGSAFNLTGPPGSAQIESLFENYYKSIKQGFVPT AFDGRSDYEGFILKGIPAGGVFTGAESLKTEEQARLFGGQAGVALDANYHAKGD NMTNLNHKAFLINSRATAFAVATYANNLSSIPPRNATVVKRESMKWTKREEPHT HGADTGCFASRVKE NB41870-WO-PCT[3] SEQ ID NO:24 neutral endoprotease; PRT; Bacillus amyloliquefaciens: MGLGKKLSVAVAASFMSLTISLPGVQAAENPQLKENLTNFVPKHSLVQSELPSVS DKAIKQYLKQNGKVFKGNPSERLKLIDQTTDDLGYKHFRYVPVVNGVPVKDSQ VIIHVDKSNNVYAINGELNNDVSAKTANSKKLSANQALDHAYKAIGKSPEAVSN GTVANKNKAELKAAATKDGKYRLAYDVTIRYIEPEPANWEVTVDAETGKILKK QNKVEHAATTGTGTTLKGKTVSLNISSESGKYVLRDLSKPTGTQIITYDLQNREY NLPGTLVSSTTNQFTTSSQRAAVDAHYNLGKVYDYFYQKFNRNSYDNKGGKIVS SVHYGSRYNNAAWIGDQMIYGDGDGSFFSPLSGSMDVTAHEMTHGVTQETANL NYENQPGALNESFSDVFGYFNDTEDWDIGEDITVSQPALRSLSNPTKYGQPDNFK NYKNLPNTDAGDYGGVHTNSGIPNKAAYNTITKIGVNKAEQIYYRALTVYLTPSS TFKDAKAALIQSARDLYGSQDAASVEAAWNAVGL

[0003] NB41870-WO-PCT[3] References Robert-Peillard, F., E. P. Barco, M. Ciulu, C. Demelas, F. Théraulaz, J.-L. Boudenne and B. Coulomb (2017). "High throughput determination of ammonium and primary amine compounds in environmental and food samples." Microchemical Journal 133: 216-221. Schlichtherle-Cerny, H. and R. Amadò (2002). "Analysis of Taste-Active Compounds in an Enzymatic Hydrolysate of Deamidated Wheat Gluten." Journal of Agricultural and Food Chemistry 50(6): 1515-1522. WO2016210395 A1: Aminopeptidase for protein hydrolysates

Claims

NB41870-WO-PCT[3] What is claimed is:

1. A method for producing a hydrolysate enriched in glutamate comprising adding a glutaminase comprising an isolated polypeptide having at least 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof to a starting material comprising glutamine and incubating for a sufficient time to produce the hydrolysate.

2. The method of claim 1 wherein the isolated polypeptide has at least 80% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

3. The method of claim 2 wherein the isolated polypeptide has at least 85% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

4. The method of claim 3 wherein the isolated polypeptide has at least 90% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

5. The method of claim 4 wherein the isolated polypeptide has at least 95% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

6. The method of claim 5 wherein the isolated polypeptide has at least 98% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13,NB41870-WO-PCT[3] SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

7. The method of claim 6 wherein the isolated polypeptide has at least 99% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

8. The method of claim 7 wherein the isolated polypeptide has 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

9. The method of any of claims 1 to 8 wherein the step of incubating is conducted at a temperature of around 40 to 70°C.

10. The method of claim 10 wherein the step of incubating is conducted at a temperature of around 45 to 65°C.

11. The method of claim 11 wherein the step of incubating is conducted at a temperature around 50 to 60°C.

12. The method of claim 11 wherein the step of incubating is conducted at a temperature around 55°C.

13. The method of any of claims 1 to 12 wherein said starting material is a plant protein.

14. The method of claim 13 wherein said plant protein is selected from the group consisting of pea, soy, fava, gluten, and oat.

15. The method of any of claims 1 to 14 further comprising adding an exo-peptidase.NB41870-WO-PCT[3] 16. The method of claim 15 wherein said exo-peptidase comprises an isolated polypeptide having at least 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:23 or an exopeptidase active fragment thereof.

17. The method of any of claims 1 to 16 further comprising adding an endo-peptidase.

18. The method of claim 17 wherein said endopeptidase comprises an isolated polypeptide having at least 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:22 or an endopeptidase active fragment thereof or SEQ ID NO:24 or an endopeptidase active fragment thereof.

19. A protein hydrolysate enriched in glutamate produced according to any of claims 1 to 18.

20. Use of a protein hydrolysate according to claim 19 to produce a broth, a meat product or a cheese.

21. A method of providing umami flavor to a plant-based meat alternative comprising the steps of: a. providing a batter comprising a plant protein; b. combining the batter with a glutaminase enzyme comprising an isolated polypeptide having at least 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof; c. incubating the batter at a temperature of about 40 to 70°C; and d. cooking the batter to provide the plant-based meat alternative.

22. The method of claim 21 wherein the isolated polypeptide has at least 80% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.NB41870-WO-PCT[3] 23. The method of claim 22 wherein the isolated polypeptide has at least 85% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

24. The method of claim 23 wherein the isolated polypeptide has at least 90% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

25. The method of claim 24 wherein the isolated polypeptide has at least 95% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

26. The method of claim 25 wherein the isolated polypeptide has at least 98% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

27. The method of claim 26 wherein the isolated polypeptide has at least 99% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

28. The method of claim 27 wherein the isolated polypeptide has 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.NB41870-WO-PCT[3] 29. The method of any of claims 21 to 28 wherein the step of incubating is conducted at a temperature of around 40 to 70°C.

30. The method of claim 29 wherein the step of incubating is conducted at a temperature of around 45 to 65°C.

31. The method of claim 30 wherein the step of incubating is conducted at a temperature around 50 to 60°C.

32. The method of claim 31 wherein the step of incubating is conducted at a temperature around 55°C.

33. The method of any of claims 21 to 33 wherein the plant protein is selected from the group consisting of pea, soy, fava, gluten, and oat.

34. The method of any of claims 21 to 33 further comprising adding an exo-peptidase at step a and / or step b.

35. The method of claim 34 wherein said exo-peptidase comprises an isolated polypeptide having 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:23 or a proteolytically active fragment thereof.

36. The method of any of claims 21 to 35 further comprising adding an endo-peptidase at step a and / or step b.

37. The method of claim 36 wherein said endo-peptidase comprises an isolated polypeptide having 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:22 or SEQ ID NO:24 or an exopeptidase active fragment thereof.NB41870-WO-PCT[3] 38. A recombinant glutaminase comprising an isolated polypeptide having at least 70, 75, 80, 85, 90, 95, 98, 99 or 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof to a starting material comprising glutamine and incubating for a sufficient time to produce the hydrolysate.

39. The recombinant glutaminase of claim 38 wherein the isolated polypeptide has at least 80% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

40. The recombinant glutaminase of claim 39 wherein the isolated polypeptide has at least 85% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

41. The recombinant glutaminase of claim 40 wherein the isolated polypeptide has at least 90% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

42. The recombinant glutaminase of claim 41 wherein the isolated polypeptide has at least 95% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

43. The recombinant glutaminase of claim 42 wherein the isolated polypeptide has at least 98% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.NB41870-WO-PCT[3] 44. The recombinant glutaminase of claim 43 wherein the isolated polypeptide has at least 99% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

45. The recombinant glutaminase of claim 44 wherein the isolated polypeptide has 100% sequence identity to SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:7, SEQ ID NO:10, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17 or SEQ ID NO:19 or a glutaminase active fragment thereof.

46. A glutaminase preparation comprising the glutaminase enzyme of any of claims 38 to 45 wherein said preparation has low endo-protease activity.

47. The glutaminase preparation of claim 46 comprising a liquid formulation, a dry formulation or a cell fermentate.

48. An isolated polynucleotide comprising a nucleic acid sequence which encodes the isolated polypeptide of any of claims 38 to 45.

49. A nucleic acid construct comprising the polynucleotide of claim 48 operably linked to one or more control sequences that direct the production of the polypeptide in a suitable expression host.

50. A recombinant expression vector comprising the nucleic acid construct of claim 49.

51. A recombinant host cell comprising the nucleic acid construct of claim 49 or the vector of claim 50.

52. A method for producing the recombinant glutaminase of any of claims 38 to 45 comprising cultivating the recombinant host cell of claim 51, to produce a supernatant and / or cells comprising the polypeptide; and recovering the polypeptide.NB41870-WO-PCT[3] 53. A recombinant glutaminase produced by the method of claim 52.

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