TW202221137A - Production of fucosylated lactose structures by a cell - Google Patents

Abstract

The present invention is in the technical field of synthetic biology and metabolic engineering. The present invention describes methods for the production of 2',3-difucosyllactose as well as the purification of said 2',3-difucosyllactose from the mixture. The present invention describes methods for the production of a mixture of 2',3-difucosyllactose, 2'-fucosyllactose and/or 3-fucosyllactose as well as the purification of any one of said 2'-fucosyllactose, 3-fucosyllactose and 2',3-difucosyllactose from the mixture. The present invention also describes a method for the production of 2',3-difucosyllactose with a purity of at least 80% based on the total amount of 2'-fucosyllactose, 3-fucosyllactose and 2',3-difucosyllactose produced. Furthermore, the present invention is in the technical field of fermentation of metabolically engineered cells. The present invention describes a cell metabolically engineered for production of 2',3-difucosyllactose or a mixture of 2',3-difucosyllactose, 2'-fucosyllactose and/or 3-fucosyllactose.

Description

Production of fucosylated lactose structures by cells

The present invention is in the technical field of synthetic biology and metabolic engineering. The present invention describes methods for producing 2',3-difucosyllactose and purifying the 2',3-difucosyllactose from mixtures. The present invention describes a mixture for the production of 2',3-difucosyllactose, 2'-fucosyllactose and/or 3-fucosyllactose and the purification of the 2'-fucosyllactose from the mixture A method of any of glycosyl lactose, 3-fucosyllactose, and 2',3-difucosyllactose. The present invention also describes a method for the production of a compound having a purity of at least 80% based on the total amount of 2'-fucosyllactose, 3-fucosyllactose and 2',3-difucosyllactose produced. A method for 2',3-difucosyllactose. Furthermore, the present invention is in the technical field of fermentation of metabolically engineered cells. The present invention describes a metabolically engineered for the production of 2',3-difucosyllactose, or 2',3-difucosyllactose, 2'-fucosyllactose and/or 3 - cells of a mixture of fucosyllactose.

Oligosaccharides, often present in sugar-bound forms with proteins and lipids, are involved in many biological phenomena such as differentiation, development and biorecognition processes associated with the initiation and progression of fertilization, embryogenesis, inflammation, cancer metastasis and host pathogen adhesion. Oligosaccharides can also be present in body fluids and human milk as unconjugated glycans, where oligosaccharides also regulate important developmental and immune processes (Bode, Early Hum. Dev. 1-4 (2015); Reily et al., Nat. Rev. . Nephrol. 15, 346-366 (2019); Varki, Glycobiology 27, 3-49 (2017)). Fucosylated lactose containing 2'-fucosyllactose (2'FL), 3-fucosyllactose (3-FL) and 2',3-difucosyllactose (DiFL) are structurally active It affects human health, as these lactose structures are described in several processes, such as protection by preventing infection of the intestinal system and by improving intestinal barrier and immune regulation. 2',3-Difucosyllactose is reported to reduce infections in the intestinal system by inhibiting the adhesion of pathogenic bacteria. 2',3-Difucosyllactose is also reported to actively support gut health by selectively stimulating bifidobacteria. Owing to the broad functional spectrum of 2'-fucosyllactose, 3-fucosyllactose and 2',3-difucosyllactose, oligosaccharide mixtures have great scientific and commercial value. However, the availability of mixtures of 2'-fucosyllactose, 3-fucosyllactose and 2',3-difucosyllactose and purified 2',3-difucosyllactose is limited by Limitations because production relies on chemical or chemical-enzymatic synthesis or on purification from natural sources such as animal milk. Chemical synthesis methods are laborious and time-consuming, and these methods are difficult to scale up due to the large number of steps involved. Enzymatic routes using fucosyltransferases offer many advantages over chemical synthesis. Fucosyltransferases catalyze the transfer of fucose moieties from an activated GDP-fucose donor to a sugar or non-sugar acceptor (Coutinho et al., J. Mol. Biol. 328 (2003) 307-317). These fucosyltransferases are the source from which biotechnologists synthesize 2'-fucosyllactose, 3-fucosyllactose and 2',3-difucosyllactose and are used (chemically) Enzymatic pathways and cell-based production systems. However, the stereospecificity and regioselectivity of fucosyltransferases remain a formidable challenge. Additionally, the chemoenzymatic pathway requires in situ regeneration of the GDP-fucose donor. Cellular production of 2'-fucosyllactose, 3-fucosyllactose, and 2',3-difucosyllactose requires tight control of adequate levels of GDP-fucose in the vicinity of complementary fucosyltransferases time and space availability.

It is an object of the present invention to provide means and methods by means of which a mixture of 2'-fucosyllactose, 3-fucosyllactose and 2',3-difucosyllactose can be Efficient, time-efficient and cost-effective way and where necessary in a continuous process. It is also an object of the present invention to provide tools and methods by means of which 2',3-difucosyllactose can be produced with a purity of at least 80% based on the amount of total oligosaccharides produced. According to the present invention, this object and other objects are achieved by providing methods and cells (preferably single cell), wherein the cell is genetically modified for the production of the fucosylated lactose structures.

Surprisingly, it has now been found that by combining at least two specific fucosyltransferases and GDP-fucose as a donor for these specific fucosyltransferases, it is possible to generate 2',3 - Difucosyllactose (DiFL), or 2',3-Difucosyllactose (DiFL), 2'-fucosyllactose (2'FL) and/or 3-fucosyllactose (3-FL). Even more surprisingly, it has now been found that it is possible to produce DiFL with a purity of at least 80%, based on the total amount of oligosaccharides produced, preferably based on the total amount of 2'FL, 3-FL and DiFL produced . The present invention provides methods and cells (preferably single cells) for producing DiFL, or a mixture of DiFL, 2'FL and/or 3-FL. The methods involve the use of cell-free systems and the use of cells expressing at least two fucosyltransferases and capable of synthesizing GDP-fucose as a donor for these fucosyltransferases. The present invention also relates to a method for producing DiFL or a mixture of DiFL, 2'FL and/or 3-FL by means of a process allowing the production of the DiFL or the mixture of diFL, 2'FL and/or 3-FL The cells were cultured under conditions. The present invention further provides cells that are metabolically engineered for the production of DiFL or a mixture of DiFL, 2'FL and/or 3-FL. The present invention also provides methods for isolating at least one of the 2'FL, 3-FL and DiFL. In addition, the present invention also provides methods for obtaining DiFL having a purity of at least 80% based on the total amount of 2'FL, 3-FL and DiFL produced. definition

Words used in this specification to describe the invention and various specific examples thereof should be understood not only in the sense of their commonly defined meanings, but also to include structures specifically defined in this specification that are outside the scope of the commonly defined meanings , material or effect. Thus, if an element can be understood in the context of this specification to include more than one meaning, its use in the scope of the claim must be understood to mean from the ordinary to all possible meanings supported by this specification and the word group itself.

The various embodiments and aspects of the embodiments disclosed herein are to be understood not only in the order and context specifically described in this specification, but also in any order and any combination thereof. Whenever the situation requires, all words used in the singular will be considered to include the plural, and vice versa. Unless otherwise defined, all technical and scientific terms used herein generally have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. In general, the nomenclature used herein and the experimental procedures in cell culture, molecular genetics, organic chemistry and nucleic acid chemistry and hybridization described herein are those well known and commonly used in the art, experiments in cell culture Procedures, Molecular Genetics, Organic Chemistry and Nucleic Acid Chemistry and Hybridization. Nucleic acid and peptide synthesis is performed using standard techniques. Generally, purification steps are carried out according to the manufacturer's instructions.

In this specification, specific examples of the present invention have been disclosed, and although specific terms are used, these terms are used in a descriptive sense only and not for the purpose of limiting the scope of the present invention, which is set forth in the following patent applications in the range. It must be understood that the specific examples described have been set forth for purposes of illustration only, and should not be construed as limiting the invention. It will be apparent to those skilled in the art that changes, other specific examples, improvements, details and uses can be made in accordance with the letter and spirit of the invention herein and within the scope of the invention, which is limited only by the scope of the patent application, according to Patent law including the doctrine of equivalents is explained. In the following claims, reference characters used to designate claims steps are provided for ease of description only, and such reference characters are not intended to imply any particular order for performing the steps.

In this document and the scope of its patent application, the verb "to comprise" and its inflections are used in their non-limiting sense, meaning to include items following the word group, but not to exclude items not specifically mentioned project. Throughout the application, the verb "to comprise" may be replaced by "to consist of" or "to consist essentially of", and vice versa. In addition, the verb "consisting of" can be replaced by "consisting essentially of", meaning that a composition as defined herein may comprise additional component(s) in addition to the specifically identified component(s), the (etc.) The additional components do not alter the unique characteristics of the present invention. In addition, reference to an element by the indefinite article "a (a/an)" does not preclude the presence of more than one element unless the context clearly requires the presence of one and only one element. Therefore the indefinite article "a" usually means "at least one". Throughout this application, unless expressly stated otherwise, the articles "a and an" are preferably replaced by "at least two", more preferably "at least three" , even better by "at least four", even better by "at least five", even better by "at least six", and most Preferably replaced by "at least two".

Throughout this application, unless expressly stated otherwise, the features "synthesize", "synthesized" and "synthesis" may be associated with the features "produce", "produced", respectively " and "production" are used interchangeably.

Specific examples as identified herein may be grouped together unless otherwise indicated. All publications, patents and patent applications mentioned in this specification are incorporated herein by reference to the same extent as if each individual publication, patent or patent application was specifically and individually indicated by reference Incorporated into general. The priority applications, including EP20190198, EP20190200 and EP20190204 in their entirety, are also incorporated herein by reference to the same extent as if specifically and individually indicated that such priority applications are incorporated by reference.

According to the present invention, the term "polynucleotide" generally refers to any polyribonucleotide or polydeoxyribonucleotide that may be unmodified RNA or DNA or modified RNA or DNA. "Polynucleotide" includes, but is not limited to, single- and double-stranded DNA; DNA that is a mixture of single- and double-stranded regions or a mixture of single-, double- and triple-stranded regions; single- and double-stranded RNA; RNA that is a mixture of stranded and double-stranded regions; hybrid molecules comprising DNA and RNA that can be single-stranded or more commonly double-stranded or triple-stranded regions, or a mixture of single-stranded and double-stranded regions. Additionally, as used herein, the term "polynucleotide" refers to a three-stranded region comprising RNA or DNA or both RNA and DNA. Strands in such regions can be from the same molecule or from different molecules. The regions may include all of one or more of the molecules, but more generally refer to regions having only some of the molecules. One of the molecules of the triple helix region is often an oligonucleotide. As used herein, the term "polynucleotide" also includes DNA or RNA as described above containing one or more modified bases. Thus, DNA or RNA having a backbone modified for stability or for other reasons is a "polynucleotide" according to the present invention. Furthermore, DNA or RNA comprising uncommon bases (such as inosine) or modified bases (such as tritylated bases) should be understood to be covered by the term "polynucleotide". It will be appreciated that various modifications have been made to DNA and RNA for many suitable purposes known to those of ordinary skill in the art. The term "polynucleotide" as used herein encompasses such chemically, enzymatically or metabolically modified forms of polynucleotides, as well as DNA and RNA specific to viruses and cells, including, for example, simple and complex cells the chemical form. The term "polynucleotide" also encompasses short polynucleotides commonly referred to as oligonucleotides.

"Polypeptide" refers to any peptide or protein comprising two or more amino acids joined to each other by peptide bonds or modified peptide bonds. "Polypeptide" refers to both short chains (often referred to as peptides, oligopeptides and oligomers) and long chains (often referred to as proteins). Polypeptides may contain amino acids other than the 20 gene-encoded amino acids. "Polypeptide" includes polypeptides modified by natural processes, such as processing and other post-translational modifications, as well as by chemical modification techniques. Such modifications are well described in the basic text and in more detailed monographs, as well as in the extensive research literature, and are well known to those of ordinary skill in the art. Modifications of the same type may be present to the same or varying degrees at several sites in a given polypeptide. Furthermore, a given polypeptide may contain various types of modifications. Modifications can occur anywhere in a polypeptide, including the peptide backbone, amino acid side chains, and amino or carboxyl termini. Modifications include, for example: acetylation, acetylation, ADP-ribosylation, acylation, covalent attachment of flavin, covalent attachment of blood matrix moieties, covalent attachment of nucleotides or nucleotide derivatives, lipids or Covalent attachment of lipid derivatives, covalent attachment of phosphatidylinositol, cross-linking, cyclization, disulfide bond formation, demethylation, formation of covalent cross-links, formation of pyroglutamic acid, methylation , γ-carboxylation, glycosylation, GPI anchor formation, hydroxylation, iodination, methylation, myristylation, oxidation, proteolytic processing, phosphorylation, prenylation, racemization, lipid linkage , sulfation, gamma-carboxylation of glutamic acid residues, hydroxylation and ADP-ribosylation, selenylation, transfer RNA-mediated addition of amino acids to proteins (such as spermidine) and ubiquitination. Polypeptides can be branched or cyclic with or without branching. Cyclic, branched, and branched cyclic polypeptides can result from post-translational natural processes, and can also be prepared by fully synthetic methods.

As used herein, the term "polynucleotide encoding a polypeptide" encompasses polynucleotides comprising sequences encoding a polypeptide of the invention. The term also encompasses polynucleotides that include single contiguous or non-contiguous regions encoding polypeptides (eg, interspersed with integrated phage or insert sequences or edits) and other regions that may also contain coding and/or non-coding sequences.

"Isolated" means altered "by the hand of man" from its natural state, that is, if it exists in nature, it is altered or removed or both from its original environment. For example, a polynucleotide or polypeptide that naturally occurs in a living organism is not "isolated," but the same polynucleotide or polypeptide that is separated from coexisting materials in its natural state is "isolated," as the term is used herein. used in. Similarly, the term "synthetic" sequence as used herein means any sequence that has been synthetically produced and not isolated directly from a natural source. The term "synthetic" as used herein means any synthetically produced sequence and not isolated directly from a natural source.

As used herein in reference to a cell or host cell, the terms "recombinant" or "transgenic" or "genetically modified" are used interchangeably and indicate that the cell replicates a heterologous nucleic acid, or Expressed by a heterologous nucleic acid (ie, a sequence "foreign to said cell" or a sequence "foreign to said location or environment in said cell") Encoded peptide or protein. Such cells are described as being transformed by, or by introducing at least one heterologous or exogenous gene. Metabolically engineered or recombinant or transgenic or genetically modified cells may contain genes not found in the native (non-recombinant) form of the cell. Recombinant cells may also contain genes found in the cell in its native form, wherein the gene has been modified by artificial means and reintroduced into the cell. These terms also encompass cells containing nucleic acid endogenous to the cell that has been modified or whose expression or activity has been modified without removing the nucleic acid from the cell; such modifications include by gene replacement, promoter replacement ; site-specific mutations; and related art-obtained modifications. Accordingly, a "recombinant polypeptide" is a polypeptide produced by recombinant cells. As used herein, a "heterologous sequence" or "heterologous nucleic acid" is derived from a source foreign to a particular cell (eg, from a different species), or from the same source if it is from the same source A sequence or nucleic acid that has been modified from its original form or position in the genome. Thus, a heterologous nucleic acid operably linked to a promoter is from a source different from the source from which the promoter was derived, or if from the same source, modified from its original form or location in the gene body. Heterologous sequences can be stably introduced into the genome of a host microbial cell, eg, by transfection, transformation, conjugation or transduction, wherein techniques are applied depending on the cell and the sequence to be introduced. Various techniques are known to those of ordinary skill in the art and are disclosed, for example, in Sambrook et al., Molecular Cloning: A Laboratory Manual, 2nd Edition, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (1989). The term "mutant" cell or microorganism as used in the context of the present invention refers to a genetically modified cell or microorganism.

In the context of the present invention, the term "endogenous" refers to anything that is a natural part of the cell and is present at its natural location in the cell's chromosomes and that behaves in contrast to the natural control mechanisms acting on its expression Control unaltered polynucleotide, polypeptide or protein sequences. The term "exogenous" refers to any polynucleotide, polypeptide that is derived from the outside of the cell under study and not from a natural part of the cell or in its natural location in the cell's chromosome or plastid or protein sequence.

When used in reference to a polynucleotide, gene, nucleic acid, polypeptide or enzyme, the term "heterologous" refers to a polynucleotide from or derived from a source other than the host organism species , gene, nucleic acid, polypeptide or enzyme. In contrast, "homologous" polynucleotides, genes, nucleic acids, polypeptides or enzymes are used herein to refer to polynucleotides, genes, nucleic acids, polypeptides or enzymes that are derived from the host organism species. When referring to gene regulatory sequences or helper nucleic acid sequences (eg, promoters, 5' untranslated regions, 3' untranslated regions, poly A addition sequences, intron sequences, splice sites, ribosome binding sites, internal ribosome entry sequences, gene body homology regions, recombination sites, etc.), "heterologous" means that a regulatory or auxiliary sequence is not associated with the regulatory or auxiliary nucleic acid sequence in a construct, gene body , chromosomes or episomal bodies with which the genes are naturally associated. Accordingly, a promoter operably linked to a gene that is not operably linked to a promoter in its natural state (ie, in the form of the gene body of a non-genetically engineered organism) is referred to herein as a "heterologous promoter" )" even though the promoter may be derived from the same species (or in some cases, the same organism) as the gene to which it is linked.

The term "modified activity" of a protein or enzyme relates to a change in the activity of a protein or enzyme compared to the wild-type (ie, native) activity of the protein or enzyme. The modified activity can be the activity of the protein or enzyme that is eliminated, attenuated, reduced or delayed compared to the wild-type activity of the protein or enzyme, but can also be the activity of the protein or enzyme compared to the wild-type activity of the protein or enzyme. The accelerated or enhanced activity of an enzyme. The modified activity of a protein or enzyme is obtained by modified expression of the protein or enzyme or by the expression of a modified (ie, mutated) form of the protein or enzyme. The modified activity of the enzyme further relates to the modification of the apparent Michaelis constant Km and/or the apparent maximum velocity (Vmax) of the enzyme.

The term "modified expression" of a gene refers to changes in the expression compared to the wild-type expression of the gene at any stage of the production process of the encoded protein. The modified expression is lower or higher than the wild type, wherein in the case of an endogenous gene, the term "higher expression" is also defined as "overexpression" of that gene, Or in the case of a heterologous gene not present in the wild-type strain, it is defined as "expression". Lower expression or reduced expression is obtained by means of common well-known techniques of those of ordinary skill in the art (such as the use of siRNA, CrispR, CrispRi, riboswitches, recombineering, homologous recombination, ssDNA mutagenesis, RNAi, miRNA, asRNA, mutant gene, knockout gene, transposon mutagenesis, ...), these techniques are used to render the gene less capable (ie, statistically significantly "less capable" compared to a functional wild-type gene) or completely A gene cannot be altered in such a way that a functional end product is produced, such as by knocking out the gene. The term "riboswitch" as used herein is defined as a portion of a messenger RNA that folds into an intricate structure that blocks expression by interfering with translation. Binding of effector molecules induces conformational changes allowing post-transcriptional regulatory expression. The gene of interest is then altered in such a way that lower performance is obtained as described above, also by altering transcription units, promoters, untranslated regions, ribosome binding sites, Shine Dalgarno sequences or transcription terminators for lower performance. Lower expression or reduced expression can be achieved, for example, by mutating one or more base pairs in the promoter sequence or completely changing the promoter sequence to a persistent promoter with lower expression strength compared to wild type or An inducible promoter that causes regulated expression or a repressible promoter that causes regulated expression can be obtained. Overexpression or expression is obtained by means of common well-known techniques to those of ordinary skill in the art (such as the use of artificial transcription factors, redesign of promoter sequences, ribosome engineering, introduction or reintroduction of expression modules at eukaryotic , using high-copy-count plastids), in which the gene is part of an "expression cassette" concerning the presence of promoter sequences, non-translated region sequences (containing ribosome-binding sequences, summer Indargano or Kozak sequences), coding sequences, and optionally translation terminators, and any sequence that results in the expression of a functionally active protein. The manifestations are persistent or accommodative.

The term "constitutive expression" is defined as being free under certain growth conditions from transcription factors other than subunits of RNA polymerase (eg, bacterial delta factors such as ^σ70 , ^σ54 , or related σ factors; and Expression of the yeast mitochondrial RNA polymerase-specific factor (MTF1) regulation by co-association with the RNA polymerase core enzyme. Non-limiting examples of such transcription factors are CRP, LacI, ArcA, Cra, IclR in E. coli, or Aft2p, Crzlp, Skn7 in Saccharomyces cerevisiae , or B. subtilis ( B. DeoR, GntR, Fur in subtilis ). These transcription factors bind to specific sequences and can block or enhance performance under certain growth conditions. RNA polymerase is the catalytic mechanism for the synthesis of RNA from DNA templates. RNA polymerase binds specific sequences to initiate transcription, eg, via delta factor in prokaryotic hosts or via MTF1 in yeast. Persistent expression provides a constant amount of expression without the need for induction or inhibition.

The term "regulated expression" is defined as an expression that is regulated by transcription factors other than subunits of RNA polymerase (eg, bacterial delta factor) under certain growth conditions. Examples of such transcription factors are described above. Typically performance modulation is obtained by means of an inducer or inhibitor such as (but not limited to) IPTG, arabinose, rhamnose, fucose, allo-lactose, or pH changes, or temperature changes, or carbon depletion or receptors or resulting products or chemical inhibition.

The term "expression by a natural inducer" is defined as a facultative or regulatory expression of a gene that is expressed only under certain natural conditions in the host (eg, an organism during parturition or during lactation), As a response to environmental changes (e.g. including, but not limited to, hormones, heat, cold, pH changes, light, oxidative or osmotic stress/signaling), or depending on the location of the developmental stage or the cell cycle of the host cell, Including (but not limited to) apoptosis and autophagy.

The term "inducible expression upon chemical treatment" is defined as a facultative or regulated expression of a gene that is expressed only upon treatment with a chemical inducer or inhibitor, wherein the inducer and inhibitor comprise ( but not limited to) alcohols (eg ethanol, methanol), carbohydrates (eg glucose, galactose, glycerol, lactose, arabinose, rhamnose, fucose, allolactose), metal ions (eg aluminium, copper, zinc) ), nitrogen, phosphate, IPTG, acetate, formate, xylene.

The term "control sequence" refers to a sequence recognized by the cellular transcription and translation system that allows transcription and translation of a polynucleotide sequence into a polypeptide. Such DNA sequences are thus necessary for the expression of the operably linked coding sequence in a particular cell or organism. Such control sequences can be, but are not limited to, promoter sequences, ribosome binding sequences, Schyndalgarno sequences, Kozak sequences, transcription terminator sequences. Control sequences suitable for use in prokaryotes include, for example, promoters, optional operator sequences, and ribosome binding sites. Eukaryotic cells are known to utilize promoters, polyadenylation signals and enhancers. The DNA of a presequence or a secretory leader sequence is operably linked to the DNA of a polypeptide if it appears to be involved in the secretion of a preprotein; a promoter or enhancer is operably linked to a coding sequence if it affects the transcription of the sequence or operably linked to a coding sequence if the ribosome binding site affects transcription of the sequence; or operably linked to a coding sequence if the ribosome binding site is positioned to facilitate translation . These control sequences may additionally be via inducible promoters or via induction or repression by external chemicals such as, but not limited to, IPTG, arabinose, lactose, allo-lactose, rhamnose or fucose The gene circuit controls the transcription or translation of the polynucleotide into a polypeptide.

In general, "operably linked" means that the DNA sequences being linked are contiguous and, in the case of a secretory leader, contiguous and in reading phase. However, enhancers need not be contiguous. The term "wild type" refers to the genetic or phenotypic condition commonly known as it occurs in nature.

The term "modified expression of a protein" as used herein refers to i) a higher expression or overexpression of an endogenous protein, ii) a heterologous protein, as compared to a wild-type (ie, native) protein expression of the protein or iii) expression and/or overexpression of a variant protein with higher activity.

As used herein, the term "mammary cell" generally refers to mammary epithelial cells, mammary epithelial luminal cells, or mammalian epithelial alveolar cells, or any combination thereof. As used herein, the term "mammary-like cell" generally refers to cells having a phenotype/genotype that is similar (or substantially similar) to natural breast cells, but derived from sources other than mammary cells. Such mammary gland-like cells can be engineered to remove at least one undesired genetic component and/or include at least one predetermined genetic construct characteristic of mammary gland cells. Non-limiting examples of mammary-like cells can include mammary epithelioid cells, mammary epithelial lumen-like cells, non-mammary cells exhibiting one or more characteristics of cells of the mammary cell lineage, or any combination thereof. Other non-limiting examples of mammary-like cells may include cells having a phenotype similar (or substantially similar) to native breast cells, or more specifically, similar (or substantially similar) to native mammary epithelial cells. Cells having a phenotype or exhibiting at least one characteristic similar (or substantially similar) to native mammary cells or mammary epithelial cells can include cells exhibiting native or having been engineered to be capable of expressing at least one milk component (e.g., derived from mammary glands). cell lineage or non-mammary cell lineage).

As used herein, the term "non-mammary cell" can generally include cells of any non-mammary lineage. In the context of the present invention, a non-mammary gland cell can be any mammalian cell capable of being engineered to express at least one milk component. Non-limiting examples of such non-mammary cells include hepatocytes, blood cells, kidney cells, umbilical cord blood cells, epithelial cells, epidermal cells, muscle cells, fibroblasts, mesenchymal cells, or any combination thereof. In some cases, molecular biology and genome editing techniques can be engineered to eliminate, silence or attenuate numerous genes simultaneously.

Throughout this application, unless expressly stated otherwise, the expressions "capable of...<verb>" and "capable to...<verb>" are preferred Substitute ground with the active voice of the verb, and vice versa. For example, the expression "capable of expressing" is preferably replaced with "express", and vice versa, ie "capable of expressing" is preferably replaced with "capable of expressing".

As used herein, the term "variant" is a polynucleotide or polypeptide that differs from a reference polynucleotide or polypeptide, respectively, but retains essential properties. A typical variant of a polynucleotide differs in nucleotide sequence from another reference polynucleotide. Changes in the nucleotide sequence of the variant may or may not alter the amino acid sequence of the polypeptide encoded by the reference polynucleotide. As discussed below, nucleotide changes may result in amino acid substitutions, additions, deletions, fusions and truncations in the polypeptide encoded by the reference sequence. A typical variant of a polypeptide differs in amino acid sequence from another reference polypeptide. Often, the differences are limited such that the sequence of the reference polypeptide and the variant are generally quite similar and identical in many regions. A variant and a reference polypeptide can differ in amino acid sequence by one or more substitutions, additions, deletions, in any combination. A substituted or inserted amino acid residue may or may not be a residue encoded by the genetic code. Variants of polynucleotides or polypeptides can be naturally-occurring, such as dual gene variants, or they can be variants not known to be naturally-occurring. Non-naturally occurring variants of polynucleotides and polypeptides can be prepared by mutagenesis techniques, by direct synthesis, and by other recombinant methods known to those of ordinary skill in the art.

As used herein, the term "derivative" of a polypeptide is one that can contain deletions, additions or substitutions of amino acid residues within the amino acid sequence of the polypeptide, but which result in silent changes, thereby producing functionally equivalent Polypeptide of polypeptide. Amino acid substitutions can be made based on similarity in polarity, charge, solubility, hydrophobicity, hydrophilicity, and/or amphiphilic properties of the residues involved. Examples of non-polar (hydrophobic) amino acids include alanine, leucine, isoleucine, valine, proline, phenylalanine, tryptophan, and methionine; planar neutral amines Base acids include glycine, serine, threonine, cysteine, tyrosine, aspartamine, and glutamic acid; positively charged (basic) amino acids include arginine, amino acids and histidine; and negatively charged (acidic) amino acids include aspartic acid and glutamic acid. In the context of the present invention, a derivative polypeptide as used herein refers to a polypeptide capable of exhibiting substantially similar in vitro and/or in vivo activities as the original polypeptide, such as by a variety of criteria including, but not limited to, enzymes activity), and which can be differentially modified during or after translation. In addition, non-classical amino acids or chemical amino acid analogs can be introduced into the original polypeptide sequence in substitution or addition form.

In some embodiments, the present invention contemplates making functional variants by modifying the structure of an enzyme as used in the present invention. Variants can be created by amino acid substitutions, deletions, additions, or combinations thereof. For example, it is reasonable to expect that independent substitution of leucine with isoleucine or valine, independent substitution of aspartic acid with glutamic acid, independent substitution of threonine with serine, or independent substitution of amine groups Similar substitution of an acid with a structurally related amino acid (eg, conservative mutation) does not have a significant effect on the biological activity of the resulting molecule. Conservative substitutions are those that occur within a family of amino acids whose side chains are related. Whether changes in the amino acid sequence of the polypeptides of the invention result in functional homologues can readily be determined by assessing the ability of the variant polypeptide to generate a response in cells in a manner similar to that of the wild-type polypeptide, in the context of the present invention compared to Provides better yield, productivity and/or growth rate in cells without the variant.

The term "functional homolog" as used herein describes those molecules that share sequence similarity (in other words, homology) and also share at least one functional characteristic (such as biochemical activity) (Altenhoff et al ., PLoS Comput. Biol. 8 (2012) e1002514). Functional homologues will typically produce the same characteristics to a similar, but not necessarily the same, degree. Functionally homologous proteins produce the same characteristics, wherein the quantitative measure produced by one homologue is at least 10% of the quantitative measure produced by the other; more typically, at least 20% of the quantitative measure produced by the original molecule , between about 30% and about 40%; for example, between about 50% and about 60%; between about 70% and about 80%; or between about 90% and about 95%; % and about 100%, or greater than 100%. Thus, where the molecule has enzymatic activity, the functional homologue will have the enzymatic activity percentages listed above compared to the original enzyme. In the case of a molecule that is a DNA binding molecule (eg, a polypeptide), the homologue will have binding affinity above the recited percentages compared to the original molecule, as measured by the weight of the binding molecule.

Functional homologs and reference polypeptides can be naturally occurring polypeptides, and sequence similarity can be due to convergent or divergent evolutionary events. Functional homologs are sometimes referred to as heterologs, where "ortholog" refers to a homologous gene or protein that is the functional equivalent of a referenced gene or protein in another species.

Heterologous genes are homologous genes in different species that arise by vertical transmission of a single gene from the last common ancestor, where the genes and their major functions are conserved. Homologous genes are genes inherited in two species by a common ancestor.

When used with reference to an amino acid or nucleotide/nucleic acid sequence from a given species, the term "heterolog" refers to the same amino acid or nucleotide/nucleic acid sequence from a different species. It will be understood that two sequences are xenologs to each other when they are derived from a common ancestral sequence through linear transmission, and/or are otherwise closely related both in terms of their sequence and their biological function. Heterologs will typically have a high degree of sequence identity but may not (and usually will not) share 100% sequence identity.

A homologous gene is a homologous gene produced by a gene duplication event. Homologous genes usually belong to the same species, but this is not required. Homologs can be split into in-paralogs (homologous pairs that appear after the speciation event) and out-paralogs (the same species that appear before the speciation event) homologous pair). Between species, an exolog is a pair of homologs that exists between two organisms due to replication prior to speciation. Within a species, an exolog is a pair of homologs that exist in the same organism but whose replication event occurs after speciation. Homologs typically have the same or similar function. Functional homologues can be identified by analysis of nucleotide and polypeptide sequence alignments. For example, performing a query on a database of nucleotide or polypeptide sequences can identify homologues of the polypeptide of interest, such as, for example, biomass-modulating polypeptides, glycosyltransferases, proteins or membranes involved in nucleotide-activated sugar synthesis transport protein. Sequence analysis may involve BLAST, Reciprocal BLAST, or Reciprocal BLAST against non-redundant databases using the amino acid sequences of biomass-modulating polypeptides, glycosyltransferases, proteins involved in nucleotide-activated sugar synthesis, or membrane transporters, respectively, as reference sequences. PSI-BLAST analysis. In some cases, the amino acid sequence is deduced from the nucleotide sequence. Typically, those polypeptides in the database with more than 40% sequence identity are candidates for further assessment as to their suitability as biomass-modulating polypeptides, glycosyltransferases, proteins involved in nucleotide-activated sugar synthesis, or membrane transport proteins, respectively. thing. Amino acid sequence similarity allows for conservative amino acid substitutions such as substitution of one hydrophobic residue by another hydrophobic residue or substitution of one polar residue by another polar residue or substitution of one acidic amino acid by another acidic amine base acid substitution or substitution of one basic amino acid with another basic amino acid, etc. Preferably, by conservative substitution, is meant a combination of, such as glycine by alanine, and vice versa; valine, isoleucine, and leucine by methionine, and Vice versa; aspartic acid is replaced by glutamic acid and vice versa; aspartic acid is replaced by glutamic acid and vice versa; serine is replaced by threonine and vice versa However; lysine is substituted by arginine, and vice versa; cysteine is substituted by methionine, and vice versa; and phenylalanine and tyrosine are substituted by tryptophan, and vice versa The same is true. If desired, manual inspection of such candidates can be performed in order to limit the number of candidates to be further evaluated. Manual inspection can be performed by selecting candidates that appear to have domains (eg, conserved functional domains) present in productivity-modulating polypeptides.

In terms of polynucleotides, "Fragment" refers to any portion of a clone or polynucleotide molecule, especially a portion of a polynucleotide that retains the functional characteristics of a full-length polynucleotide molecule. Useful fragments include oligonucleotides and polynucleotides that can be used in hybridization or amplification techniques or to modulate replication, transcription or translation. "Polynucleotide fragment" refers to any subsequence of a polynucleotide of SEQ ID NO (or Genbank NO.), which typically comprises the nucleotide sequence of any of the polynucleotide sequences provided herein. At least about 9, 10, 11, 12 contiguous nucleotides or consist thereof, eg, at least about 30 nucleotides or at least about 50 nucleotides. Exemplary fragments may additionally or alternatively include a region comprising, consisting essentially of, or consisting of a conserved family domain encoding a polypeptide. Exemplary fragments may additionally or alternatively include fragments comprising conserved domains of polypeptides. Therefore, a fragment of a polynucleotide SEQ ID NO (or Genbank NO.) preferably means a nucleotide sequence comprising or consisting of the polynucleotide SEQ ID NO (or Genbank NO.), of which no more than 200 , 150, 100, 50 or 25 consecutive nucleotide deletions, preferably no more than 50 consecutive nucleotide deletions, and the nucleotide sequence retention can be assessed by those of ordinary skill in the art through routine experimentation Available functional characteristics (eg, activity) of full-length polynucleotide molecules. Alternatively, a fragment of a polynucleotide SEQ ID NO (or Genbank NO.) preferably means a nucleotide sequence comprising a quantified contiguous nucleus from the polynucleotide SEQ ID NO (or Genbank NO.) nucleotides or consisting of, and wherein the certain amount of contiguous nucleotides is at least 50.0%, 60.0%, 70.0%, 80.0%, 81.0% of the full length of the polynucleotide SEQ ID NO (or Genbank NO.) , 82.0 %, 83.0 %, 84.0 %, 85.0 %, 86.0 %, 87.0 %, 88.0 %, 89.0 %, 90.0 %, 91.0 %, 92.0 %, 93.0 %, 94.0 %, 95.0 %, 95.5 %, 96.0 %, 96.5 %, 97.0%, 97.5%, 98.0%, 98.5%, 99.0%, 99.5%, 100%, preferably at least 80.0%, more preferably at least 87.0%, even better at least 90.0%, even better at least 95.0%, most Preferably at least 97% and retain the useful functional characteristics (eg, activity) of the full-length polynucleotide molecule. Therefore, a fragment of a polynucleotide SEQ ID NO (or Genbank NO.) preferably means a nucleotide sequence comprising the polynucleotide SEQ ID NO (or Genbank NO.) or consisting of the same, wherein a certain amount of Consecutive nucleotide deletions and wherein the amount does not exceed 50.0%, 40.0%, 30.0% of the full length of the polynucleotide SEQ ID NO (or Genbank NO.), preferably not more than the polynucleotide SEQ ID NO (or Genbank NO.) or Genbank NO.) of the full length of 20.0%, 15.0%, 10.0%, 9.0%, 8.0%, 7.0%, 6.0%, 5.0%, 4.5%, 4.0%, 3.5%, 3.0%, 2.5%, 2.0%, 1.5%, 1.0%, 0.5%, better not more than 15.0%, even better not more than 10.0%, even better not more than 5.0%, best not more than 2.5%, and wherein the fragment retention can be determined by the relevant technical field available functional characteristics (eg, activity) of full-length polynucleotide molecules as routinely assessed by those of ordinary skill in .

Throughout the application, polynucleotide sequences may be represented by SEQ ID NOs or, alternatively, by GenBank NOs. Accordingly, unless expressly stated otherwise, the terms "polynucleotide SEQ ID NO" and "polynucleotide GenBank NO." are used interchangeably.

Fragments may additionally or alternatively include subsequences of polypeptides and protein molecules, or subsequences of polypeptides. In some cases, a fragment or domain is a subsequence of a polypeptide that performs at least one biological function of the intact polypeptide in substantially the same manner as the intact polypeptide, preferably to a similar extent. A "subsequence of the polypeptide" as defined herein refers to a sequence derived from contiguous amino acid residues of a polypeptide. For example, a polypeptide fragment can contain identifiable structural motifs or functional domains, such as DNA binding sites or domains that bind to DNA promoter regions, activation domains, or protein-protein interaction domains, and can initiate transcription. Fragments can vary in size from as few as 3 amino acid residues to the full length of a complete polypeptide, eg, at least about 20 amino acid residues in length, eg, at least about 30 amino acid residues in length. Thus, a fragment of a polypeptide SEQ ID NO (or UniProt ID or Genbank NO.) preferably means a polypeptide sequence comprising or consisting of the polypeptide SEQ ID NO (or UniProt ID or Genbank NO.), wherein no more than 80 , 60, 50, 40, 30, 20, or 15 consecutive amino acid residues are deleted, preferably no more than 40 consecutive amino acid residues are deleted, and it depends on those skilled in the art. Routinely assessed intact polypeptides preferably perform at least one biological function of the intact polypeptide to a similar or greater extent in substantially the same manner. Alternatively, a fragment of a polypeptide SEQ ID NO (or UniProt ID or Genbank NO.) preferably means a polypeptide sequence comprising a quantification of contiguous amino acids from the polypeptide SEQ ID NO (or UniProt ID or Genbank NO.) residues or consisting of, and wherein the certain amount of contiguous amino acid residues is at least 50.0%, 60.0%, 70.0%, 80.0%, 81.0 %, 82.0 %, 83.0 %, 84.0 %, 85.0 %, 86.0 %, 87.0 %, 88.0 %, 89.0 %, 90.0 %, 91.0 %, 92.0 %, 93.0 %, 94.0 %, 95.0 %, 95.5 %, 96.0 % , 96.5%, 97.0%, 97.5%, 98.0%, 98.5%, 99.0%, 99.5%, 100%, preferably at least 80.0%, more preferably at least 87.0%, even better at least 90.0%, even better at least 95.0% , optimally at least 97.0%, and which preferably perform at least one biological function of the intact polypeptide to a similar or greater extent in substantially the same manner as the intact polypeptide can be routinely assessed by those of ordinary skill in the art. Thus, a fragment of a polypeptide SEQ ID NO (or UniProt ID or Genbank NO.) preferably means a polypeptide sequence comprising or consisting of the polypeptide SEQ ID NO (or UniProt ID or Genbank NO.), wherein a certain amount of Consecutive amino acid residues are deleted and wherein the amount does not exceed 50.0%, 40.0%, 30.0% of the full length of the polypeptide SEQ ID NO (or UniProt ID or Genbank NO.), preferably not more than the polypeptide SEQ ID NO (or UniProt ID or Genbank NO.) 20.0%, 15.0%, 10.0%, 9.0%, 8.0%, 7.0%, 6.0%, 5.0%, 4.5%, 4.0%, 3.5%, 3.0%, 2.5%, 2.0 %, 1.5%, 1.0%, 0.5%, better not more than 15.0%, even better not more than 10.0%, even better not more than 5.0%, and best not more than 2.5%, and its dependence can be achieved by the relevant technology Intact polypeptides routinely assessed by those of ordinary skill in the art preferably perform at least one biological function of the intact polypeptide in substantially the same manner, preferably to a similar or greater extent.

Throughout the application, polypeptide sequences may be represented by SEQ ID NO or alternatively by UniProt ID or GenBank NO. Thus, unless expressly stated otherwise, the terms "polypeptide SEQ ID NO" and "polypeptide Uniprot ID" and "polypeptide GenBank NO. (polypeptide GenBank NO.)" are used interchangeably.

Preferably, a fragment of a polypeptide is a functional fragment derived from a polypeptide that preferably possesses at least one property or activity of the polypeptide to a similar or greater extent. Functional fragments may, for example, include functional or conserved domains of polypeptides. It will be appreciated that a polypeptide or fragment thereof may have conservative amino acid substitutions that have substantially no effect on the activity of the polypeptide. By conservative substitution, it means that one hydrophobic amino acid is substituted with another hydrophobic amino acid or one polar amino acid is substituted with another polar amino acid or one acidic amino acid is substituted with another acidic amino acid Or one basic amino acid is substituted by another basic amino acid, etc. Preferably, by conservative substitution, is meant a combination of, such as glycine by alanine, and vice versa; valine, isoleucine, and leucine by methionine, and Vice versa; aspartic acid is replaced by glutamic acid and vice versa; aspartic acid is replaced by glutamic acid and vice versa; serine is replaced by threonine and vice versa However; lysine is substituted by arginine, and vice versa; cysteine is substituted by methionine, and vice versa; and phenylalanine and tyrosine are substituted by tryptophan, and vice versa The same is true. Domains can be identified, for example, by Pfam (El-Gebali et al., Nucleic Acids Res. 47 (2019) D427-D432) or the Conserved Domain Database (CDD) (https://www.ncbi.nlm.nih. gov/cdd) (Lu et al, Nucleic Acids Res. 48 (2020) D265-D268) name qualification. The content of each database is fixed and does not change with each release. When the content of a particular database changes, this particular database receives a new release version with a new release date. All release versions of each database and their corresponding release dates and specific content as noted on such specific release dates are available and known to those skilled in the art. The PFAM database (https://pfam.xfam.org/) used in this article is Pfam version 33.1 released on June 11, 2020. Protein sequence information and functional information can be provided by comprehensive sources of protein sequence and annotation data, such as, for example, the Universal Protein Resource (UniProt) (www.uniprot.org) (Nucleic Acids Res. 2021, 49(D1), D480-D489). UniProt includes a specialized and well-managed protein database called the UniProt Knowledge Base (UniProtKB), as well as the UniProt Reference Sequence Set (UniRef) and the UniProt Archive (UniParc). UniProt identifiers (UniProt IDs) are unique to each protein present in the database. UniProt ID as used herein is the UniProt ID in the UniProt database version on May 05, 2021. The NIH Gene Sequence Database (https://www.ncbi.nlm.nih.gov/genbank/) (Nucleic Acids Res. 2013, 41(D1), D36) is used in this paper as it exists on May 05, 2021 -D42) The respective Genbank Accession Number (GenBank No.) in the version refers to proteins that do not have UniProt IDs.

The term "fucosyltransferase" as used herein refers to an enzyme capable of catalyzing the transfer of a fucose moiety from an activated GDP-fucose donor molecule to a specific acceptor molecule, thereby forming a glycosidic bond. Fucosyltransferases belonging to the superclass of glycosyltransferases have been described and classified into different sequence-based families (Campbell et al., Biochem. J. 326, 929-939 (1997) ) and available on the CAZy (Carbohydrate Active Enzymes) website (www.cazy.org). Fucosyltransferase includes α-1,2-fucosyltransferase, α-1,3-fucosyltransferase, α-1,4-fucosyltransferase and α-1, 6-fucosyltransferase, which catalyzes the transfer of Fuc residues from GDP-Fuc to glycan acceptors via an alpha-glycosidic bond. Fucosyltransferases can be found but are not limited to the GT10, GT11, GT23, GT65 and GT68 CAZy families.

The terms "alpha-1,2-fucosyltransferase", "alpha 1,2 fucosyltransferase" as used in the present invention , "2-fucosyltransferase (2-fucosyltransferase)", "α-1,2-fucosyltransferase (α-1,2-fucosyltransferase)", "α1,2fucosyltransferase" Transferase (α 1,2 fucosyltransferase)", "2 fucosyltransferase (2 fucosyltransferase)", "2-FT" or "2FT" are used interchangeably and refer to the catalysis of fucose from the donor GDP-L- Glycosyltransferase that transfers fucose to acceptor molecules in α-1,2-bonds. As used herein, the acceptor molecule may be lactose, resulting in the formation of 2'-fucosyllactose. Additionally, the acceptor molecule used herein can also be 3-fucosyllactose, resulting in the formation of 2',3-difucosyllactose.

The terms "alpha-1,3-fucosyltransferase", "alpha 1,3 fucosyltransferase" as used in the present invention , "3-fucosyltransferase (3-fucosyltransferase)", "α-1,3-fucosyltransferase (α-1,3-fucosyltransferase)", "α1,3fucosyltransferase" Transferase (α 1,3 fucosyltransferase)", "3 fucosyltransferase (3 fucosyltransferase)", "3-FT" or "3FT" are used interchangeably and refer to the catalysis of fucose from the donor GDP-L- A glycosyltransferase that transfers fucose to acceptor molecules in α-1,3-bonds. As used herein, the acceptor molecule can be lactose, resulting in the formation of 3-fucosyllactose. Additionally, the acceptor molecule used herein may also be 2'-fucosyllactose, resulting in the formation of 2',3-difucosyllactose.

The terms "2' fucosyllactose", "2'-fucosyllactose", "α-1,2-fucosyl" as used in the present invention Lactose (α-1,2-fucosyllactose)", "α-1,2-fucosyllactose (α-1,2-fucosyllactose)", "α-1,2-fucosyllactose (α-1,2- fucosyllactose)", "α 1,2 fucosyllactose (α 1,2 fucosyllactose)", "Galβ-4(Fucα1-2)Glc", "2FL" or "2'FL" are used interchangeably and refer to borrowed Product obtained by catalyzing the transfer of a fucose residue from GDP-L-fucose to lactose in an α-1,2-bond by α-1,2-fucosyltransferase. As used in the present invention, the terms "difucosyllactose", "di-fucosyllactose", "lactodifucotetraose", "2', 3-difucosyllactose (2',3-difucosyllactose)", "2',3 difucosyllactose (2',3 difucosyllactose)", "α-2',3-difucose Glucosyl lactose (α-2', 3-difucosyllactose)", "α 2', 3 difucosyllactose (α 2', 3 difucosyllactose)", "Fucα1-2Galβ 1-4(Fucα1-3)Glc" , "DFLac", "2',3 diFL", "DFL", "DiFL" or "diFL" are used interchangeably.

The terms "3-fucosyllactose (3-fucosyllactose)", "α-1,3-fucosyllactose (alpha-1,3-fucosyllactose)", "α1,3 Fucosyllactose (alpha 1,3 fucosyllactose)", "alpha-1,3-fucosyllactose (alpha-1,3-fucosyllactose)", "alpha 1,3 fucosyllactose (alpha 1 ,3 fucosyllactose)", "Galβ-4(Fucα1-3)Glc", "3FL" or "3-FL" are used interchangeably and refer to the conversion of rock by catalyzing α-1,3-fucosyltransferase A product obtained by the transfer of a halose residue from GDP-L-fucose to lactose in an α-1,3-bond.

The term "monosaccharide" as used herein refers to sugars containing only one simple sugar that cannot be broken down by hydrolysis into simpler sugars and that contain one or more hydroxyl groups per molecule. As used herein, monosaccharide refers to glucose, galactose and fucose. As used herein, the term "disaccharide" refers to a sugar composed of two monosaccharide units such as, for example, lactose (Gal-bl,4-Glc).

The term "Oligosaccharide" as used herein refers to a sugar polymer containing three to four simple sugars, ie, monosaccharides. Monosaccharides as used herein are reducing sugars and include glucose, galactose and fucose. A glycoside between two sugar units can be represented, for example, as 1,2, 1->2, or (1-2), used interchangeably herein. For example, the terms "Gal-b1,4-Glc", "β-Gal-(1->4)-Glc", "Galβ1-4-Glc" and "Gal-b(1-4)-Glc" It has the same meaning, that is, the carbon-1 of galactose (Gal) is linked with the β-glycosidic bond of carbon-4 of glucose (Glc). Each monosaccharide may be in a cyclic form (eg, piperanose in the form of furanose). Linkages between individual monosaccharide units include α 1->2, α 1->3, and β 1->4. The oligosaccharides of the invention are constructed from the disaccharide lactose (Gal-beta1,4-Glc) which is monofucosylated on the Gal residues in α1->2 to yield 2'-fucosyllactose, or Monofucosylated on Glc residues in α1->3 linkages to yield 3-fucosyllactose, or doublefucosylated to carry Gal residues with α1->2 linkages and a fucose residue on the Glc residue linked with an α1->3 linkage to produce 2',3-difucosyllactose. As used herein and as generally understood in the current state of the art, a "fucosylated oligosaccharide" is an oligosaccharide bearing a fucose residue. As used herein, "fucosylated oligosaccharide" refers to 2'-fucosyllactose, 3-fucosyllactose, and 2',3-difucosyllactose. As used herein, the term "fucosylated lactose structure" also refers to 2'-fucosyllactose, 3-fucosyllactose, and 2',3-difucosyllactose.

A "fucosylation pathway" as used herein is a biochemical pathway consisting of enzymes and their respective genes, mannose-6-phosphate isomerase, phosphomannose mutase, mannose- Guanosyl 1-phosphate transferase, GDP-mannose 4,6-dehydratase, GDP-L-fucose synthase and/or the reuse pathway L-fucose kinase/GDP-fucose pyrophosphate fucosylases, and fucosyltransferases that produce alpha 1,2- or alpha 1,3-fucosylated oligosaccharides.

Terms "mannose-6-phosphate isomerase", "phosphomannose isomerase", "mannose phosphate isomerase", "phosphate "phosphohexoisomerase", "phosphomannoisomerase", "phosphomannose-isomerase", "phosphohexomutase", " D-mannose-6-phosphate ketol-isomerase" and "manA" are used interchangeably and refer to the reversible conversion of D-fructose 6-phosphate to D - Enzyme of mannose 6-phosphate.

Terms "phosphomannomutase", "mannose phosphomutase", "phosphomannose mutase", "D-mannose 1,6-phosphate mutase" D-mannose 1,6-phosphomutase" and "manB" are used interchangeably and refer to an enzyme that catalyzes the reversible conversion of D-mannose 6-phosphate to D-mannose 1-phosphate.

Terms "mannose-1-phosphate guanylyltransferase", "GTP-mannose-1-phosphate guanylyltransferase" , "phosphomannose isomerase-guanosine 5'-diphosphate-D-mannose pyrophosphorylase (phosphomannose isomerase-guanosine 5'-diphospho-D-mannose pyrophosphorylase; PIM-GMP)", "GDP-mannose "GDP-mannose pyrophosphorylase", "guanosine 5'-diphospho-D-mannose pyrophosphorylase", "guanosine 5'-diphospho-D-mannose pyrophosphorylase" Sugar pyrophosphorylase (guanosine diphosphomannose pyrophosphorylase), "guanosine triphosphate-mannose 1-phosphate guanylyltransferase", "mannose 1-phosphate guanylyltransferase" "mannose 1-phosphate guanylyltransferase (guanosine triphosphate)" and "manC" are used interchangeably and refer to the use of GTP to convert D-mannose-1-phosphate to GDP-mannose and Diphosphate enzyme.

Terms "GDP-mannose 4,6-dehydratase", "guanosine 5'-diphosphate-D-mannose oxidoreductase" mannose oxidoreductase)", "guanosine diphosphomannose oxidoreductase", "guanosine diphosphomannose 4,6-dehydratase", "GDP-D -Mannose dehydratase (GDP-D-mannose dehydratase)", "GDP-D-mannose 4,6-dehydratase (GDP-D-mannose 4,6-dehydratase)", "GDP-mannose 4,6 -Hydrogen-dissociating enzyme (GDP-mannose 4,6-hydro-lyase)", "GDP-mannose 4,6-hydro-lysing enzyme (forming GDP-4-dehydro-6-deoxy-D-mannose) ) (GDP-mannose 4,6-hydro-lyase (GDP-4-dehydro-6-deoxy-D-mannose-forming))” and “gmd” are used interchangeably and refer to both GDP-rhamnose and GDP- The enzyme that forms the first step in the biosynthesis of fucose.

Terms "GDP-L-fucose synthase", "GDP-4-keto-6-deoxy-D-mannose-3,5-epimerase-4-reduction" Enzyme (GDP-4-keto-6-deoxy-D-mannose-3,5-epimerase-4-reductase)", "GDP-L-fucose:NADP+4-oxidoreductase (3,5-reductase)" Isomerizing) (GDP-L-fucose:NADP+ 4-oxidoreductase (3,5-epimerizing))" and "fcl" are used interchangeably and refer to the enzyme that forms the second step in the biosynthesis of GDP-fucose .

Terms "L-fucokinase/GDP-fucose pyrophosphorylase (L-fucokinase/GDP-fucose pyrophosphorylase)", "L-fucose kinase/L-fucose-1-P guanosine L-fucokinase (L-fucokinase/L-fucose-1-P guanylyltransferase), "GDP-fucose pyrophosphorylase", "GDP-L-fucose pyrophosphorylase (GDP-L-fucose pyrophosphorylase)" GDP-L-fucose pyrophosphorylase)" and "fkp" are used interchangeably and refer to an enzyme that catalyzes the conversion of L-fucose-1-phosphate to GDP-fucose using GTP.

The term "purified" refers to a substance that is substantially or substantially free of components that interfere with the activity of a biomolecule. With respect to cells, carbohydrates, nucleic acids, and polypeptides, the term "purified" refers to a material that is substantially or substantially free of components that typically accompany the material in the form in which it is found in its natural state. Typically, a purified saccharide, oligosaccharide, protein or nucleic acid of the invention is at least about 50%, 55%, 60%, 65%, 70%, 75%, 80% or 85% pure, usually at least about 90%, 91 %, 92 %, 93 %, 94 %, 95 %, 96 %, 97 %, 98 % or 99 % pure, as measured by band intensities based on silver-stained gels or other methods for determining purity Measurement. Purity or homogeneity can be indicated by a variety of means well known in the art, such as polyacrylamide gel electrophoresis of protein or nucleic acid samples followed by observation after staining. For some purposes, high resolution and the use of HPLC or similar means for purification will be required. For oligosaccharides, purity can be determined using methods such as, but not limited to, thin layer chromatography, gas chromatography, NMR, HPLC, capillary electrophoresis, or mass spectrometry.

The terms "identical" or "identical" or "identical" in the context of two or more nucleic acid or polypeptide sequences when compared and aligned for maximum correspondence, as measured using a sequence comparison algorithm or by visual inspection "Percent identity" or "% identity" refers to two or more sequences or subsequences that are identical or have a specified percentage of identical amino acid residues or nucleotides. For sequence comparison, one sequence acts as a reference sequence to which test sequences are compared. When using a sequence comparison algorithm, test and reference sequences are entered into a computer, subsequence coordinates are specified if necessary, and sequence algorithm program parameters are specified. Next, the sequence comparison algorithm calculates the percent sequence identity of the test sequence relative to the reference sequence according to the specified program parameters. The percent identity can be calculated globally within the full-length sequence of the reference sequence, resulting in an overall percent identity score. Alternatively, percent identity can be calculated within a partial sequence of a reference sequence, resulting in a percent local identity score. The use of the full length of the reference sequence in a local sequence alignment yields an overall percent identity score between the test and reference sequences. The percent identity can use different algorithms such as, for example, BLAST and PSI-BLAST (Altschul et al., 1990, J Mol Biol 215:3, 403-410; Altschul et al., 1997, Nucleic Acids Res 25: 17, 3389-402) , Clustal Omega method (Sievers et al, 2011, Mol. Syst. Biol. 7:539), MatGAT method (Campanella et al, 2003, BMC Bioinformatics, 4:29) or EMBOSS Needle assay.

The Basic Local Alignment Search Tool (BLAST)) method of alignment is an algorithm provided by the National Center for Biotechnology Information (NCBI) to compare sequences using preset parameters. The program compares nucleotide or protein sequences to sequence databases and calculates statistical significance. The Position-Specific Iterative Basic Local Alignment Search Tool (PSI-BLAST) uses protein-protein BLAST (BLASTp) to align multiple sequences from sequences detected above a given score threshold Pairs derive a position-specific scoring matrix (PSSM) or distribution profile. BLAST methods can be used for pairwise or multiple sequence alignment. Pairwise sequence alignment is used to identify regions of similarity, which can indicate the functional, structural and/or evolutionary relationship between two biological sequences (protein or nucleic acid). The web interface for BLAST is available at: https://blast.ncbi.nlm.nih.gov/Blast.cgi.

Clustal Omega (Clustal W) is a multiple sequence alignment program that uses inoculated guide trees and the HMM profile-profile technique to generate alignments between three or more sequences. It produces biologically meaningful multiple sequence alignments of divergent sequences. The web interface for Clustal W is available at https://www.ebi.ac.uk/Tools/msa/clustalo/. The default parameters for multiple sequence alignment and percent protein sequence identity calculation using the Clustal W method are: enable de-alignment of input sequences: FALSE; enable mbed-like clustering bootstrap tree: TRUE; enable mbed-like clustering iteration: TRUE; ( Combined bootstrap tree/HMM) number of iterations: preset(0); max bootstrap tree iterations: preset[-1]; max HMM iterations: preset[-1]; command: compare.

The Matrix Global Alignment Tool (MatGAT) is a computer application that generates similarity/identity matrices for DNA or protein sequences without the need for pre-alignment of data. The program uses the Myers and Miller global alignment algorithm to perform a series of pairwise alignments, computes similarity and agreement, and then places the results in a distance matrix. Users can specify which type of alignment matrix (eg, BLOSUM50, BLOSUM62, and PAM250) to use for their protein sequence checks.

The EMBOSS Needle (https://galaxy-iuc.github.io/emboss-5.0-docs/needle.html) uses the Needleman-Wunsch global alignment algorithm when its entire length is considered Find the best alignment of two sequences (including gaps). The best alignment is ensured by a dynamic programming method by exploring all possible alignments and selecting the best one. The Needleman-Wunsch algorithm is a member of a class of algorithms that can compute optimal scores and alignments in the order of mn steps (where 'n' and 'm' are the lengths of the two sequences). Gap opening penalty (default 10.0) is the score deducted when a gap is created. The default values assume you are using the EBLOSUM62 matrix for protein sequences. A gap extension (default 0.5) penalty is added to the standard gap penalty for each base or residue in the gap. This is the way to punish long gaps.

As used herein, a polypeptide having an amino acid sequence of at least 80% sequence identity to the full-length sequence of the reference polypeptide sequence is understood to be 80%, 81%, 82%, 83% to the full-length amino acid sequence of the reference polypeptide sequence , 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 91.50%, 92.00%, 92.50%, 93.00%, 93.50%, 94.00%, 94.50%, 95.00%, 95.50 100% sequence identity. Throughout the application, unless expressly specified otherwise, amino acid sequences containing at least 80% sequence identity to the full-length amino acid sequence (or nucleotide sequence) of the reference polypeptide (or nucleotide sequence) ( or nucleotide sequence) polypeptide (or DNA sequence)/consisting of amino acids with at least 80% sequence identity with the full-length amino acid sequence (or nucleotide sequence) of the reference polypeptide (or nucleotide sequence) Polypeptide (or DNA sequence) composition of the sequence (or nucleotide sequence)/amines with at least 80% sequence identity to the full-length amino acid sequence (or nucleotide sequence) of the reference polypeptide (or nucleotide sequence) The polypeptide (or DNA sequence) of the nucleotide sequence (or nucleotide sequence), usually indicated by SEQ ID NO or UniProt ID or Genbank NO., preferably has at least 85.0%, 90.0%, 91.0%, 92.0% with the full-length reference sequence %, 93.0%, 94.0%, 95.0%, 96.0%, 97.0%, 98.0% or 99.0%, more preferably at least 85.0%, even more preferably at least 90.0%, most preferably at least 95.0% sequence identity.

For purposes of the present invention, percent identity was determined using MatGAT2.01 (Campanella et al., 2003, BMC Bioinformatics 4:29). The following preset parameters for proteins were used: (1) gap cost presence: 12 and extension: 2; (2) the matrix used was BLOSUM65. In a preferred embodiment, sequence identity is calculated based on a given SEQ ID NO, ie, the full-length sequence or a portion of the reference sequence. A portion thereof preferably means at least 50.0%, 60.0%, 70.0%, 80.0%, 90.0% or 95.0% of the complete reference sequence.

The term "cultivation" refers to the medium in which the cells are cultured or fermented, the cells themselves, and the oligosaccharides produced by the cells in whole culture broth, ie, both inside (intracellular) and outside (extracellular) cells.

The term "membrane transporter protein" as used herein refers to a protein that is part of or interacts with a cell membrane and controls the flow of molecules and information across a cell. Thus, membrane proteins are involved in transport, whether they are imported into or exported from cells.

Such membrane transporters may be transporters as defined by the Transporter Classification Database, P-P-bond hydrolysis driven transporters, β-Barrel Porins , paratransporters, putative transporters, and phosphotransfer-driven group translocation proteins, a database operated and managed by the Saier Lab Bioinformatics Group via www.tcdb.org and provides a functional and phylogenetic classification of membrane transporters . This transporter classification database details the IUBMB-approved comprehensive classification system for membrane transporters, called the Transporter Classification (TC) system. TCDB taxonomy searches as described herein are defined based on TCDB.org as published on June 17, 2019.

Transporter is the collective name for uniporters, symporters, and antiporters that utilize vector-mediated processes (Saier et al., Nucleic Acids Res. 44 (2016) D372-D379). It belongs to the class of electrochemical potential-driven transport proteins and is also known as a secondary carrier-type enhancer. Membrane transporters are included in this category when using carrier-mediated processes to catalyze uniporters when a single species is transported by facilitated diffusion or in a membrane potential-dependent process (if the solute is charged) ; catalytic antiporters when two or more species are transported in opposite directions in a tightly coupled process, not coupled to a direct energy form other than chemoosmotic energy; and/or when two or more species are transported in opposite directions When multiple species are transported together in the same direction in a tightly coupled process and are not coupled to a direct energy form other than chemoosmotic energy, catalyzing cotransport proteins, which are all secondary carriers (Forrest et al., Biochim. Biophys. Acta 1807 (2011) 167-188). These systems are usually stereospecific. Solute: Solute reverse transport is a typical feature of secondary carriers. The dynamic association of transport proteins and enzymes produces a functional membrane transport metabolite that imports substrates typically obtained from the extracellular compartment directly into their cellular metabolism (Moraes and Reithmeier, Biochim. Biophys. Acta 1818 ( 2012), 2687-2706). Solutes transported via this transporter system include, but are not limited to, cations, organic anions, inorganic anions, nucleosides, amino acids, polyols, phosphorylated glycolytic intermediates, osmotic agents, chelating iron. The term polyol means an alcohol containing multiple hydroxyl groups. For example, glycerol, sorbitol or mannitol.

A membrane transporter is included in P-P-bond hydrolysis-driven transport if it hydrolyzes the diphosphate bond of an inorganic pyrophosphate, ATP, or another nucleoside triphosphate to drive active uptake and/or extrusion of a solute or solutes in the class of proteins (Saier et al, Nucleic Acids Res. 44 (2016) D372-D379). Membrane transporters may or may not be temporarily phosphorylated, but substrates are not. Substrates transported by classes of transporters driven by hydrolysis of P-P-bonds include, but are not limited to, cations, heavy metals, beta-glucans, UDP-glucose, lipopolysaccharides, teichoic acid.

Beta-barrel porin membrane transport proteins form transmembrane pores that generally allow solutes to pass through the membrane in an energy-independent manner. The transmembrane portion of these proteins consists solely of beta strands, forming a beta-barrel (Saier et al., Nucleic Acids Res. 44 (2016) D372-D379). These porin-type proteins are found in the outer membrane of Gram-negative bacteria, mitochondria, plastids and possibly acid-fast Gram-positive bacteria. Solutes transported through these β-barrel porins include, but are not limited to, nucleosides, raffinose, glucose, β-glucosides, oligosaccharides.

Auxiliary transport proteins are defined as proteins that facilitate transport across one or more biological membranes but are not themselves directly involved in transport. These membrane transporters consistently function in conjunction with one or more existing transport systems such as (but not limited to) outer membrane factor (OMF), polysaccharide (PST) transporter, ATP-binding cassette (ATP) -binding cassette; ABC) type transport protein. It may provide functions associated with energy coupling for transport, play a structural role in complex formation, provide biogenic or stabilizing functions or regulatory functions (Saier et al, Nucleic Acids Res. 44 (2016) D372-D379). Examples of helper transport proteins include, but are not limited to, the family of polysaccharide copolymerases involved in the transport of polysaccharides, the family of membrane fusion proteins involved in the transport of bacteriocins and chemical toxins.

Putative transporters comprise the following families which will be classified elsewhere when the transport function of the members is confirmed, or eliminated from the transporter classification system if the proposed transport function proves ineffective. These families include one or more members for which transport functions have been proposed, but evidence for such functions has not been convincing (Saier et al, Nucleic Acids Res. 44 (2016) D372-D379). Examples of putative transport proteins as classified in this group under the TCDB system published on June 17, 2019 include, but are not limited to, copper transport proteins.

Phosphate transfer-driven group translocation protein is also known as bacterial phosphoenolpyruvate:sugar phosphotransferase system (phosphotransferase system; PTS) PEP-dependent phosphonium group transfer-driven group translocation protein. The reaction products derived from extracellular sugars are cytoplasmic phosphate sugars. Enzymatic components that catalyze the phosphorylation of sugars are superimposed on the transport process in a tightly coupled process. The PTS system is involved in many different aspects, including regulation and chemotaxis, biofilm formation and pathogenesis (Lengeler, J. Mol. Microbiol. Biotechnol. 25 (2015) 79-93; Saier, J. Mol. Microbiol. Biotechnol. 25 (2015) 73-78). As in the TCDB system published on June 17, 2019, the membrane transport protein family within the phosphate transfer-driven group translocation proteins includes proteins related to glucose-glucoside, fructose-mannitol, lactose-N,N'-di A PTS system associated with the transport of acetylchitobiose-beta-glucoside, glucitol, galactitol, mannose-fructose-sorbose and ascorbate.

The major facilitator superfamily (MFS) is a superfamily of membrane transport proteins that catalyze uniport proteins, solute: cation (H+, but hardly Na+) symporters and/or solutes :H+ or solute:solute antiporter. Most are 400-600 amido residues in length and have 12, 14 or occasionally 24 transmembrane alpha-helical spanners (TMS), as transported by Saier Lab Bioinformatics Group (www.tcdb.org) Defined by the Protein Taxonomy Database.

"SET" or "Sugar Efflux Transporter" as used herein refers to a membrane protein of the SET family, which is a protein with the InterPRO domain IPR004750 and/or a protein belonging to the eggNOGv4.5 family ENOG410XTE9. InterPro domains can be identified by using the online tool at https://www.ebi.ac.uk/interpro/ or the standalone version of InterProScan (https://www.ebi.ac.uk/interpro/download.html) Use preset values. Identification of orthologous families in eggNOGv4.5 can be performed using the online or standalone version of eggNOG-mapperv1 (http://eggnogdb.embl.de/#/app/home).

The term "Siderophore" as used herein refers to the secondary metabolites of various microorganisms, which are primarily iron ion-specific chelators. These molecules have been classified as catecholate, hydroxamate, carboxylates and mixed types. Chelaterin is generally synthesized by a nonribosomal peptide synthetase (NRPS)-dependent pathway or an NRPS-independent pathway (NIS). The most important precursor in the NRPS-dependent chelatin biosynthesis pathway is chorismate. 2,3-DHBA can be formed from chorismate by a three-step reaction catalyzed by isochorismate synthase, isochorismate enzyme and 2,3-dihydroxybenzoate-2,3-dehydrogenase. Chelaterin can also be formed from salicylates, which are formed from isochorismates by isochorismate pyruvate lyase. When ornithine is used as a chelatin precursor, biosynthesis depends on the hydroxylation of ornithine catalyzed by L-ornithine N5-monooxygenase. In the NIS pathway, an important step in chelatin biosynthesis is N(6)-hydroxylysine synthase.

Transport proteins are required to export chelatin to the outside of the cell. So far, four superfamilies of membrane proteins have been identified in the process: the major facilitator superfamily (MFS); the multidrug/oligosaccharidyl-lipid/polysaccharide flippase superfamily (Multidrug/Oligosaccharidyl-lipid/Polysaccharide Flippase Superfamily; MOP ); the resistance, nodulation and cell division superfamily (RND); and the ABC superfamily. In general, genes involved in chelatin export are clustered with chelatin biosynthesis genes. The term "siderophore exporter" as used herein refers to such transport proteins required to export chelatin to the outside of the cell.

The ATP-binding cassette (ABC) superfamily contains uptake and efflux transport systems, and members of these two groups are usually loosely clustered together. ATP hydrolysis without protein phosphorylation provides energy for transport. There are dozens of families within the ABC superfamily, and families are generally associated with substrate specificity. Members are classified according to class 3.A.1 as defined by the Transport Protein Taxonomy Database operated by the Saier Lab Bioinformatics Group, available through www.tcdb.org and providing the functionality and systems of membrane transport proteins Classification occurs.

The term "enabled efflux" means the activity of introducing transport of solutes on the cytoplasmic membrane and/or cell wall. This transport can be achieved by introducing and/or increasing the expression of transport proteins as described in the present invention. The term "enhanced efflux" means an activity that improves the transport of solutes across the cytoplasmic membrane and/or cell wall. Transport of solutes across the cytoplasmic membrane and/or cell wall can be enhanced by introducing and/or increasing the expression of membrane transport proteins as described herein. "Expression" of a membrane transport protein is defined as "overexpression" of the gene encoding the membrane transport protein in the case of an endogenous gene; or in the case of the gene encoding the membrane transport protein being The absence of a heterologous gene in a wild-type strain or cell is defined as "expression".

The term "precursor" as used herein refers to substances that are taken up or synthesized by cells to specifically produce 2'FL, 3-FL and DiFL of the present invention. Examples of such precursors in this sense include fructose, glycerol, sucrose, mannose, fucose, GDP-fucose, mannose-6-phosphate, mannose-1-phosphate, GDP-mannose Sugar and GDP-4-dehydro-6-deoxy-α-D-mannose.

The term "acceptor" as used herein refers to a disaccharide or oligosaccharide that can be modified by a fucosyltransferase, resulting in the synthesis of 2'-fucosyllactose, 3-fucosyllactose and/or 2',3-Difucosyllactose. Acceptors as used herein include lactose, 2'-fucosyllactose, and 3-fucosyllactose. DETAILED DESCRIPTION OF THE INVENTION

According to a first aspect, the present invention provides a method for producing 2',3-difucosyllactose (DiFL). The method includes the following steps: i) Provide at least two fucosyltransferases, preferably at least one α-1,2-fucosyltransferase and at least one α-1,3-fucosyltransferase, and GDP-fucosyltransferase sugar donors, wherein the fucosyltransferases are capable of transferring fucose residues from the GDP-fucose donor to one or more acceptors, and, ii) subjecting the fucosyltransferase and GDP-fucose donor to a mixture comprising acceptor lactose and possibly one or more other acceptors on the fucosyltransferase catalyzing fucose residues Contacting under conditions that transfer from the GDP-fucose donor to the acceptor(s), thereby producing DiFL and possibly 2'FL and/or 3FL, iii) Preferably, at least one of the 2'FL, 3-FL and DiFL is separated, more preferably, all of the 2'FL, 3-FL and DiFL are separated.

In one embodiment, the present invention provides a method for producing a mixture of DiFL, 2'-fucosyllactose (2'FL) and/or 3-fucosyllactose (3-FL). The method includes the following steps: i) Provide at least two fucosyltransferases, preferably at least one α-1,2-fucosyltransferase and at least one α-1,3-fucosyltransferase, and GDP-fucosyltransferase sugar donors, wherein the fucosyltransferases are capable of transferring fucose residues from the GDP-fucose donor to one or more acceptors, and, ii) subjecting the fucosyltransferase and GDP-fucose donor to a mixture comprising acceptor lactose and possibly one or more other acceptors on the fucosyltransferase catalyzing fucose residues Contacting under conditions in which the GDP-fucose donor is transferred to the acceptor(s), iii) Preferably, at least one of the DiFL, 2'FL and/or 3-FL is isolated, more preferably, all of the DiFL, 2'FL and/or 3-FL are isolated.

In a specific example, the present invention provides a method for the production of 2'-fucosyllactose (2'FL), 3-fucosyllactose (3-FL) and 2',3-difucosyl Method for mixtures of lactose (DiFL). The method includes the following steps: i) Provide at least two fucosyltransferases, preferably at least one α-1,2-fucosyltransferase and at least one α-1,3-fucosyltransferase, and GDP-fucosyltransferase sugar donors, wherein the fucosyltransferases are capable of transferring fucose residues from the GDP-fucose donor to one or more acceptors, and, ii) subjecting the fucosyltransferase and GDP-fucose donor to a mixture comprising acceptor lactose and possibly one or more other acceptors on the fucosyltransferase catalyzing fucose residues Contacting under conditions in which the GDP-fucose donor is transferred to the acceptor(s), iii) Preferably, at least one of the 2'FL, 3-FL and DiFL is separated, more preferably, all of the 2'FL, 3-FL and DiFL are separated.

Throughout the application, unless expressly stated otherwise, the feature "at least two fucosyltranferases" is preferably characterized by "at least one alpha-1,2-fucosyltransferase and at least one fucosyltranferase" An alpha-1,3-fucosyltransferase (at least one alpha-1,2-fucosyltransferase and at least one alpha-1,3-fucosyltransferase)" substitution.

According to a second aspect, the present invention provides a metabolically engineered cell for the production of DiFL, ie, a cell that is metabolically engineered for the production of DiFL. Provided herein is a single metabolically engineered cell capable of expressing, preferably expressing at least two fucosyltransferases capable of converting fucose residues from GDP-fucose The donor is transferred to the acceptor lactose, and possibly any one or more of the acceptor 2'FL and/or 3-FL, and the cell is capable of synthesizing GDP-fucose, wherein the GDP-fucose is used donors for these fucosyltransferases.

In one embodiment, the present invention provides a metabolically engineered cell for producing a mixture of 2'FL, 3-FL, and DiFL, that is, metabolically engineered for the production of 2'FL, 3-FL, - FL and DiFL cells. Provided herein are single metabolically engineered cells capable of expressing, preferably expressing at least two fucosyltransferases, preferably at least one alpha-1,2-fucosyltransferase, and at least one alpha- 1,3-fucosyltransferases capable of transferring fucose residues from GDP-fucose donors to acceptor lactose, and possible acceptors 2'FL and/or or any one or more of 3-FL, and the cell is capable of synthesizing GDP-fucose, wherein the GDP-fucose is a donor for the fucosyltransferases. Throughout this application, unless expressly stated otherwise, "genetically modified cell" or "metabolically engineered cell" preferably means genetically modified or metabolically modified, respectively cells engineered for use in producing the DiFL (or a mixture of DiFL, 2'FL and/or 3-FL or a mixture of 2'FL, 3-FL and DiFL) according to the invention. In the context of the present invention, DiFL, a mixture of DiFL, 2'FL and/or 3-FL or a mixture of 2'FL, 3-FL and DiFL as disclosed herein preferably is not metabolically engineered in wild-type precursor cells.

In one embodiment, the present invention provides a method for producing DiFL, wherein the fucosyltransferases and the GDP-fucose are provided in a cell-free system. In this context, at least two fucosyltransferases according to the invention, a GDP-fucose donor and optionally an acceptor are provided in the reaction mixture. Throughout the application, with regard to the cell-free method according to the present invention, the feature "culture" may be replaced with a "reaction mixture".

In one embodiment, the present invention provides a method for producing a mixture of 2'FL, 3-FL, and DiFL, wherein the fucosyltransferases and the GDP-fucose are provided in a cell-free system. In this context, at least two fucosyltransferases according to the invention, a GDP-fucose donor and optionally an acceptor are provided in the reaction mixture.

In one embodiment, the present invention provides a method for the production of DiFL by cells, preferably single cells. According to the present invention, the method for producing DiFL may utilize non-metabolically engineered cells or may utilize metabolically engineered cells as disclosed herein. In a specific example, fucosyltransferase and GDP-fucose are produced by cells.

In an alternative preferred embodiment, the present invention provides a method for producing a mixture of 2'FL, 3-FL and DiFL by cells, preferably single cells. According to the present invention, the method for producing a mixture of 2'FL, 3-FL and DiFL can utilize non-metabolically engineered cells or can utilize metabolically engineered cells as disclosed herein. In a specific example, fucosyltransferase and GDP-fucose are produced by cells, and/or provided by cells, preferably individual cells. In the context of the present invention, it is understood that the DiFL and the mixture of 2'FL, 3-FL and DiFL according to the present invention are preferably synthesized intracellularly. Those of ordinary skill in the art will further appreciate that some or substantially all of the DiFL, 2'FL and/or 3-FL produced remains intracellularly and/or is excreted outside the cell via passive or active transport.

In another preferred embodiment, the present invention provides a method for producing a mixture of 2'FL, 3-FL and DiFL, wherein the total amount of 2'FL, 3-FL and DiFL produced in the mixture is based on The DiFL is present in the mixture at a purity of at least 80%, preferably at least 85%, more preferably at least 90%, even better at least 95%, as measured. A DiFL purity of at least 80% in a mixture is understood to be the amount of 80% or more of this DiFL in a mixture of 2'FL, 3-FL and DiFL, based on the ratio of 2'FL, 3-FL and DiFL produced. Measured by the total meter, including 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 95.5, 96, 96.5, 97, 97.5, 98, 98.5, 99 or 99.5% DiFL. This method to generate mixtures with >80% DiFL can be obtained by cell-free systems. Alternatively, the method to generate mixtures with >80% DiFL can be obtained with cells, preferably single cells. Preferably, the method for producing mixtures with >80% DiFL is obtained by metabolically engineered cells, preferably individual metabolically engineered cells.

In another preferred embodiment, the present invention provides a method for producing DiFL, wherein the DiFL is at least 80%, preferably at least 85%, as measured by the total amount of oligosaccharides in the culture or reaction mixture. %, preferably at least 90%, even better at least 95% pure. DiFL purity of at least 80% should be understood as the amount of this DiFL of 80% or more of the total amount of oligosaccharides in the culture or reaction mixture, measured by the total amount of oligosaccharides produced, including 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 95.5, 96, 96.5, 97, 97.5, 98, 98.5, 99, or 99.5% DiFL. This method to produce products with >80% DiFL can be obtained by cell-free systems. Alternatively, the method to generate mixtures with >80% DiFL can be obtained by cells. Preferably, the method for producing mixtures with >80% DiFL is obtained by metabolically engineered cells.

In another preferred embodiment of the method of the present invention, precursors and/or acceptors for the synthesis of any of the 2'FL, 3-FL and DiFL are used to feed the culture or reaction mixture. Precursors and/or acceptors that can be fed into the culture or reaction mixture for the synthesis of any of the 2'FL, 3-FL and DiFL comprise lactose, 2'FL, 3-FL, fucose , glucose, galactose, sucrose, glycerol. In a specific example, the acceptor system is selected from the list comprising lactose, 2'FL and 3-FL. In a specific example, the other acceptor(s) are selected from a list comprising 2'FL and 3-FL.

In a specific example, the culture or reaction mixture is fed only with lactose.

In a specific example, the cell is capable of producing lactose intracellularly.

Lactose can be produced, preferably intracellularly, by cells (eg, by metabolism of cells and/or after metabolic engineering of cells for this purpose as known to those of ordinary skill in the art), or It can be added to cells that can import the lactose via passive or active transport. The production of lactose by cells can be obtained by expressing N-acetylglucosamine β-1,4-galactosyltransferase and UDP-glucose 4-epimerase. More preferably, the cells are modified for enhanced lactose production. The modification may be any one or more selected from the group comprising: overexpression of N-acetylglucosamine β-1,4-galactosyltransferase, overexpression of UDP-glucose 4-epimerase Excessive performance.

In a preferred embodiment of the method and/or cell of the present invention, the cell using lactose as a receptor in the glycosylation reaction (ie fucosylation according to the present invention) preferably has for uptake from culture Lactose transport protein. More preferably, the cells are optimized for lactose uptake. The optimization may be an overexpression of a lactose transporter, such as a lactose permease from Escherichia coli or Kluyveromyces lactis . Preferably, the cell continuously expresses the lactose permease. Lactose can be added at the beginning of the culture, or it can be added when sufficient biomass has formed during the growth phase of the culture, i.e., the DiFL (or mixture of 2'FL, 3-FL and DiFL) production phase (by The addition of lactose to the culture begins) separate from the growth phase. In a preferred embodiment, lactose is added at the beginning and/or during the culture, ie the growth phase and the production phase are not separated.

In a preferred embodiment of the method and/or cell according to the present invention, the cell is resistant to lactose killing when grown in an environment where lactose is combined with one or more other carbon sources. In the context of the term "lactose killing" it is meant that the growth of cells in a medium in which lactose and another carbon source are present is stunted. In a preferred embodiment, the cells are genetically modified such that they maintain at least 50% influx of lactose without undergoing lactose kill, even at high lactose concentrations, as described in WO 2016/075243. The genetic modification includes expression and/or overexpression of exogenous and/or endogenous lactose transporter genes by a heterologous promoter that does not produce a lactose-killing phenotype, and/or modification of a gene that does not produce a lactose-killing phenotype The codon usage of the lactose transporter produces an altered representation of the lactose transporter. The contents of WO 2016/075243 are incorporated by reference in this regard.

In a specific example, the culture or reaction mixture is fed with lactose, 2'FL and 3FL. In a specific example, the culture or reaction mixture is fed with lactose and 2'FL. In a specific example, the culture or reaction mixture is fed with lactose and 3FL. It may be necessary to feed the culture or reaction mixture with 2'FL and/or 3FL to obtain a specific ratio of 2'FL, 3FL and DiFL in the mixture of 2'FL, 3FL and DiFL produced by the cells.

In a specific example, the method includes the following steps: a. providing a cell, preferably a single cell, wherein the cell expresses at least two fucosyltransferases, preferably at least one alpha-1,2-fucosyltransferase and at least one alpha-1,3-peptidase Fluorosyltransferases capable of transferring a fucose residue from a GDP-fucose donor to one or more of the acceptor lactose and possibly the other acceptor(s) , and the cell is capable of synthesizing GDP-fucose, wherein the GDP-fucose is a donor for the fucosyltransferases, and b. culturing the cells under conditions that allow expression of the fucosyltransferases and synthesis of the GDP-fucose, c. Preferably, at least one of the 2'FL, 3-FL or DiFL is isolated from the culture, more preferably all of the 2'FL, 3-FL and DiFL are isolated from the culture.

Within the scope of the present invention, permissive conditions are understood to be conditions related to physical or chemical parameters including, but not limited to, temperature, pH, pressure, osmotic pressure and product/precursor/acceptor concentrations.

In a specific embodiment, allowable conditions may include a temperature range of 30 +/- 20 degrees Celsius, a pH range of 7 +/- 3.

In a preferred embodiment, the cells are cultured under fed-batch conditions and under conditions that allow expression of the fucosyltransferases and synthesis of the GDP-fucose.

According to a preferred embodiment of the present invention, cells as described herein are modified with one or more expression modules. These expression modules are also referred to as transcription units and comprise polynucleotides for expression of recombinant genes, including the coding gene sequence and appropriate transcriptional and/or translational control signals operably linked to the coding gene. Such control signals include promoter sequences, non-translated regions, ribosome binding sites, terminator sequences. The expression modules may contain elements for expressing a single recombinant gene (eg α-1,2-fucosyltransferase or α-1,3-fucosyltransferase according to the invention) but also Can contain elements for expressing more recombinant genes or can be organized for two or more recombinant genes (eg α-1,2-fucosyltransferase and/or α-1, 3-fucosyltransferase) in the operon structure expressed by the integration. Such polynucleotides can be produced by recombinant DNA technology using techniques well known in the art. Methods of constructing expression modules well known to those of ordinary skill in the art include, for example, in vitro recombinant DNA techniques, synthetic techniques, and in vivo gene recombination. See, eg, techniques described in: Sambrook et al. (2001) Molecular Cloning: a laboratory manual, 3rd edition, Cold Spring Harbor Laboratory Press, CSH, New York or Current Protocols in Molecular Biology, John Wiley and Sons, N.Y. (1989 and updated annually).

The performance of each of the performance modules may be continuous or generated by natural or chemical inducers, preferably the performance modules are continuous. As used herein, persistent expression is understood to mean the expression of genes that are continuously transcribed in an organism. Expression by a natural inducer is to be understood as a facultative or regulatory expression of a gene that is expressed only under certain natural conditions of the host (such as an organism during parturition or during lactation), as a response to environmental changes (such as including ( but not limited to) responses to hormones, heat, cold, pH changes, light, oxidative or osmotic stress/signaling), or depending on the location of the developmental stage or the cell cycle of the host cell, including but not limited to apoptosis death and autophagy. Expression by chemical inducers should be understood as sensing external chemicals (eg IPTG, arabinose, lactose, Facultative or regulatory expression of genes expressed after allolactose, rhamnose or fucose).

The expression module can be integrated into the genome of the cell or can be presented to the cell on a vector. The vector may exist in plastid, cosmid, phage, liposome or viral form stably transformed/transfected into the metabolically engineered cell. Such vectors include, inter alia, chromosomal, episomal, and virus-derived vectors, for example, vectors derived from bacterial plastids, phages, transposons, yeast episomes, insertion elements, yeast chromosomal elements, viral vectors, and vectors derived from combinations thereof , such as vectors derived from plastid and phage genetic elements such as cosmids and phages. Such vectors may contain selectable markers such as, but not limited to, antibiotic markers, auxotrophic markers, toxin-antitoxin markers, RNA sense/antisense markers. A performance system construct may contain control areas that regulate and generate performance. Generally, in this regard, any system or vector suitable for maintaining, amplifying or expressing polynucleotides and/or expressing polypeptides in a host can be used for expression. Appropriate DNA sequences can be inserted into the expression system by any of a variety of well-known and conventional techniques, such as, for example, those described in Sambrook et al., supra. For recombinant production, cells can be genetically engineered to incorporate the expression system or portions thereof or polynucleotides of the invention. Introduction of polynucleotides into cells can be accomplished by methods described in a number of standard laboratory manuals, such as Davis et al, Basic Methods in Molecular Biology, (1986) and Sambrook et al, 1989, supra.

As used herein, expression modules comprise polynucleotides for expression of at least one recombinant gene. The recombinant gene is involved in the expression of a polypeptide that functions in the synthesis of any of 2'FL, 3-FL, and DiFL; or the recombinant gene is associated with other pathways in the host cell that are not involved in 2' Synthesis of any of FL, 3-FL and DiFL. The recombinant genes encode endogenous proteins with modified expression or activity, preferably the endogenous proteins are overexpressed; or the recombinant genes encode heterogenous introductions and are expressed in the modified cells, preferably overexpressed heterologous protein. An endogenous protein can have a modified expression in cells that also express the heterologous protein.

According to a preferred embodiment of the present invention, the performance of each of the performance modules is sustained or produced by natural inducers. As used herein, persistent expression is understood to mean the expression of genes that are continuously transcribed in an organism. Expression by a natural inducer is to be understood as a facultative or regulatory expression of a gene that is expressed only under certain natural conditions of the host (such as an organism during parturition or during lactation), as a response to environmental changes (such as including ( but not limited to) responses to hormones, heat, cold, pH changes, light, oxidative or osmotic stress/signaling), or depending on the location of the developmental stage or the cell cycle of the host cell, including but not limited to apoptosis death and autophagy.

The present invention provides different types of cells for this production of DiFL or a mixture of 2'FL, 3-FL and DiFL using single cells. In a specific example, such individual cells are metabolically engineered cells. For example, the present invention provides a cell, wherein the cell expresses two fucosyltransferases and the cell synthesizes GDP-fucose, which is the fucosyl for the expression Donor of both transferases. The present invention also provides a cell, wherein the cell expresses three different fucosyltransferases and the cell synthesizes GDP-fucose, the GDP-fucose being one of all the three expressed fucosyltransferases donor. The present invention also provides a cell, wherein the cell expresses four or more different fucosyltransferases and the cell synthesizes GDP-fucose, which is the rock for all such expressed Donor of glucosyltransferase.

In one embodiment, DiFL or a mixture of 2'FL, 3-FL and DiFL is produced as a result of using at least two fucosyltransferases, preferably at least one alpha-1,2-fucose Syltransferase and at least one alpha-1,3-fucosyltransferase and a mixture of a GDP-fucose donor and a mixture comprising the acceptor lactose and possibly one or more other acceptors as described herein in a fucoid The glycosyltransferase catalyzes the transfer of fucose residues from the GDP-fucose donor to the acceptor under conditions of contact. In a specific example, the receptor used to generate the mixture of 2'FL, 3-FL and DiFL is lactose. Other receptors can be 2'FL and/or 3-FL.

In another preferred embodiment of the method and/or cell according to the present invention, the cell preferably comprises multiple copies of the same coding DNA sequence encoding a protein. In the context of the present invention, the protein may be a glycosyltransferase (eg, an alpha-1,2-fucosyltransferase or a-1,3-fucosyltransferase as described herein), Membrane protein or any other protein as disclosed herein. Throughout the application, the feature "multiple" means at least 2, preferably at least 3, more preferably at least 4, even better at least 5. These multiple copies may be integrated into the genome, may be present on the same vector/plastid and/or may be present on two or more different vectors/plastids.

In the context of the present invention, at least two fucosyltransferases are used to produce DiFL (or a mixture of 2'FL, 3-FL and DiFL) as described herein. Preferably, at least one α-1,2-fucosyltransferase and at least one α-1,3-fucosyltransferase are used to produce DiFL (or 2'FL, 3-FL) according to the invention and DiFL mixture). The DNA sequences encoding the α-1,2-fucosyltransferase and α-1,3-fucosyltransferase may be integrated into the gene body, may be present on the same vector/plastid and/or may be Present on two or more different vectors/plastids. Multiple different α-1,2-fucosyltransferases and/or α-1,3-fucosyltransferases (such as the fucosyl described herein) are used in cells or methods according to the invention transferase) are also within the scope of the present invention.

Some α-1,2-fucosyltransferases, such as Helicobacter pylori α-1,2-fucosyltransferase (SEQ ID NO 16), can additionally function as α-1 under specific conditions ,3-fucosyltransferase activity, such as H. pylori alpha-1,2-fucosyltransferase at lactose concentrations much lower than the concentration of 2'FL produced Additional modifications to 2'FL are known in the art to generate DiFL. Such α-1,2-fucosyltransferases can be used in the cells and methods according to the invention, but an α-1,3-fucosyltransferase such as α-1,3 should additionally be used - Fucosyltransferase. Preferably, the α-1,2-fucosyltransferase according to the present invention does not exert α-1,3-fucosyltransferase activity.

In a specific example, the cell-free system as described herein comprises: a. At least one alpha-1,2-fucosyltransferase that (i) comprises the polypeptide sequence of any one of SEQ ID NOs: 16 to 110, or (ii) is SEQ ID NO: 16-110 Functional homologues, variants or derivatives of any of ID NOs 16-110 with any of the alpha-1,2-fucosyltransferase polypeptides having SEQ ID NOs 16-110 The full length of which has at least 80% overall sequence identity and α-1,2-fucosyltransferase activity for any of the receptors, or (iii) is any of SEQ ID NOs 16 to 110 A functional fragment of one and having alpha-1,2-fucosyltransferase activity, or (iv) comprising a polypeptide comprising a full-length amino acid sequence having the same properties as any one of SEQ ID NOs 16-110 an amino acid sequence of at least 80% sequence identity or consisting of it and having alpha-1,2-fucosyltransferase activity, and b. At least one alpha-1,3-fucosyltransferase that (i) comprises the polypeptide sequence of any one of SEQ ID NOs: 111 to 132, or (ii) is SEQ ID NO: 111 to 132 Functional homologues, variants or derivatives of any of ID NOs 111 to 132 with any of the alpha-1,3-fucosyltransferase polypeptides having SEQ ID NOs 111 to 132 The full length of which has at least 80% overall sequence identity and α-1,3-fucosyltransferase activity for any of the receptors, or (iii) is any of SEQ ID NOs 111 to 132 A functional fragment of one and having alpha-1,3-fucosyltransferase activity, or (iv) comprising a polypeptide comprising the same full-length amino acid sequence as any one of SEQ ID NOs 111-132 an amino acid sequence of at least 80% sequence identity or consisting of it and having alpha-1,3-fucosyltransferase activity, and c. GDP-fucose, and d. Lactose and possibly at least one of these other receptor(s).

Throughout the application, features "any one of SEQ ID NOs 16 to 110" are preferably used with features "SEQ ID NOs 19 to 21, 23 to 38, 40 to 44" , 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 (any one of SEQ ID NOs 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105) ", more preferably with the feature "SEQ ID NO 19 or 25 (SEQ ID NO 19 or 25)". Alternatively, throughout the application, the feature "any one of SEQ ID NOs 16 to 110" is preferably used with the feature "any one of SEQ ID NOs 16, 17, 18 or 19" Replacement of any one of SEQ ID NOs 16, 17, 18 or 19, more preferably with the feature "SEQ ID NO 16 or 19 (SEQ ID NO 16 or 19)", even more preferably with the feature "SEQ ID NOs 16 or 19" ID NO 16" replacement.

Throughout the application, the feature "any one of SEQ ID NOs 111 to 132" is preferably used with the feature "any one of SEQ ID NOs 113 to 132" of SEQ ID NOs 113 to 132)” replacement, more preferably with the feature “any one of SEQ ID NOs 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132 of SEQ ID NOs 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132)" replacement, even better with the feature "SEQ ID NOs 113, 114, 118, 119, 120, 121 or 122 (any one of SEQ ID NOs 113, 114, 118, 119, 120, 121 or 122)" substitution, even more preferably with the feature "SEQ ID NOs 113 or 114 (SEQ ID NOs 113 or 122)" 114)" replacement, the best replacement with the feature "SEQ ID NO 114".

In a specific example, a cell as described herein expresses two fucosyltransferases, wherein at least one of the fucosyltransferases is an alpha-1,2-fucosyltransferase and At least one other of the fucosyltransferases is an alpha-1,3-fucosyltransferase.

In a specific example, cells as described herein express a. At least one alpha-1,2-fucosyltransferase, the fucosyltransferase (i) comprising the polypeptide sequence of any of the following: SEQ ID NOs: 16, 17, 18, 19 , 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44 , 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69 , 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94 , 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, or 110, or (ii) are functional homologues of any of the following, Variant or derivative: SEQ ID NO 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36 , 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61 , 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86 , 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, or 110, and with SEQ ID NOs 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109 or 110 The full length of any of the alpha-1,2-fucosyltransferase polypeptides has at least 80% overall sequence identity and is identical to lactose and possibly the other receptor(s) 2'FL and/or 3FL either has alpha-1,2-fucosyltransferase activity, or (iii) is SEQ ID NOs 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27 , 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52 , 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77 , 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102 A functional fragment of any of , 103, 104, 105, 106, 107, 108, 109, or 110 and having alpha-1,2-fucosyltransferase activity, or (iv) comprising a polypeptide comprising with SEQ ID NOs 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, Full length of any of 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109 or 110 An amino acid sequence having or consisting of an amino acid sequence having at least 80% sequence identity and having alpha-1,2-fucosyltransferase activity, and b. At least one alpha-1,3-fucosyltransferase, the fucosyltransferase (i) comprising as SEQ ID NOs: 111, 112, 113, 114, 115, 116, 117, 118, 119 , 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131 or 132. The polypeptide sequence of any one, or (ii) is SEQ ID NO 111, 112, 113, 114, Functional homologue, variant or derivative of any of 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131 or 132 substances with SEQ ID NOs 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131 or 132 The full-length of any of the α-1,3-fucosyltransferase polypeptides has at least 80% overall sequence identity and is identical to lactose and possibly the (etc.) other receptors 2'FL and/or 3FL any one of which has fucosyltransferase activity, or (iii) it is SEQ ID NOs 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124 A functional fragment of any one of , 125, 126, 127, 128, 129, 130, 131 or 132 and having alpha-1,3-fucosyltransferase activity, or (iv) comprising a polypeptide comprising with any of SEQ ID NOs 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131 or 132 The full-length amino acid sequence of one has or consists of an amino acid sequence of at least 80% sequence identity and has alpha-1,3-fucosyltransferase activity.

As used herein, one of the α-1,2-fucosyltransferases and one of the α-1,3-fucosyltransferases and a GDP-fucose donor With mixtures comprising lactose and possibly 2'FL and/or 3-FL, the fucosyltransferases catalyze the transfer of fucose residues from the GDP-fucose donor to the acceptor(s). Contact under conditions sufficient to produce a mixture of 2'FL, 3-FL and DiFL. Alternatively, two of the α-1,2-fucosyltransferases and one of the α-1,3-fucosyltransferases and a GDP-fucose donor are combined with the A mixture of lactose and possibly 2'FL and/or 3-FL under conditions under which the fucosyltransferases catalyze the transfer of fucose residues from the GDP-fucose donor to the acceptor(s) The contact is sufficient to produce DiFL or a mixture of 2'FL, 3-FL and DiFL. In addition, one of the α-1,2-fucosyltransferases and both of the α-1,3-fucosyltransferases and the GDP-fucose donor were combined with the A mixture of lactose and possibly 2'FL and/or 3-FL under conditions under which the fucosyltransferases catalyze the transfer of fucose residues from the GDP-fucose donor to the acceptor(s) The contact is sufficient to produce DiFL or a mixture of 2'FL, 3-FL and DiFL. Another alternative comprises making two or more of the alpha-1,2-fucosyltransferases and two or more of the alpha-1,3-fucosyltransferases and GDP-fucose donors with mixtures comprising lactose and possibly 2'FL and/or 3-FL where the fucosyltransferases catalyze the removal of fucose residues from the GDP-fucose donor The transfer to the acceptor(s) is contacted under conditions sufficient to produce DiFL or a mixture of 2'FL, 3-FL and DiFL.

In another embodiment of the methods and/or cells of the invention, the cells are modified in the expression or activity of at least one of the fucosyltransferases. In a preferred embodiment, the fucosyltransferase is an endogenous protein of a cell with a modified expression or activity, preferably the endogenous fucosyltransferase is overexpressed; alternatively, the fucosyltransferase Glycosyltransferases are heterologous proteins that are introduced heterogeneously and expressed in the cell, preferably overexpressed. The endogenous fucosyltransferase can have a modified expression in cells that also express the heterologous fucosyltransferase.

In an embodiment of the method and/or cell according to the invention, the cell is capable of synthesizing GDP-fuc. GDP-fucose can be provided by enzymes expressed in cells or by cellular metabolism. Such cells producing GDP-fucose may express enzymes that convert, for example, fucose added to the cell to GDP-fucose. This enzyme can be, for example, a bifunctional fucosokinase/fucose-1-phosphate guanosyltransferase, such as Fkp from Bacteroides fragilis , or a respective fucose kinase together with a respective A combination of allofucose-1-phosphate guanosyltransferases, as known from several species, including Homo sapiens, Sus scrofa and Rattus norvegicus .

Preferably, the cells are modified to produce GDP-fucose. More preferably, the cells are modified for enhanced GDP-fucose production. The modification may be any one or more selected from the group comprising: UDP-glucose: undecaprenyl-phosphoglucose-1-phosphotransferase-encoding gene deletion, GDP-L-rock Overexpression of the gene encoding alucose synthase, overexpression of the gene encoding GDP-mannose 4,6-dehydratase, overexpression of the gene encoding mannose-1-phosphate guanosyltransferase, phosphomannose mutase Overexpression of encoding genes and overexpression of genes encoding mannose-6-phosphate isomerase. Throughout the application, unless expressly stated otherwise, features "enhanced" and/or "optimized" production preferably mean modifications and/or metabolisms introduced into cells as described herein Engineering results in higher yields compared to wild-type precursor cells of the modified cells or metabolically engineered cells. For example, "enhanced GDP-fucose production" preferably means higher intracellular production of GDP-fucose in modified cells compared to wild-type precursor cells that do not contain these specific modifications .

Preferably, the cells in this context comprise a fucosylation pathway as described herein.

In a preferred embodiment of the method and/or cell of the present invention, any one of the 2'FL, 3-FL or DiFL, preferably all of the 2'FL, 3-FL or DiFL, by Translocation to the outside of the cell occurs by passive transport, ie without consuming energy from the cell by means of active transport systems.

In another preferred embodiment of the method and/or cell of the present invention, the cell is further metabolically engineered for use in i) Modified expression of endogenous membrane proteins, and/or ii) the modified activity of the endogenous membrane protein, and/or iii) Representation of homologous membrane proteins, and/or iv) expression of heterologous membrane proteins, wherein the membrane protein is involved in secreting any of the 2'FL, 3-FL and DiFL from the mixture to the outside of the cell. The cell may express one of the membrane proteins involved in the secretion of any of the 2'FL, 3-FL and DiFL from the cell to the outside of the cell. The cell may also express more than one of the membrane proteins. Any of the membrane proteins can translocate one or more of the 2'FL, 3-FL and DiFL to the outside of the cell. The cell producing DiFL or a mixture of 2'FL, 3-FL and DiFL can translocate any of the 2'FL, 3-FL and DiFL, the cell comprising passive diffusion, channels, respectively Membrane proteins, membrane transport proteins, membrane carrier proteins.

In a specific example, cells as described herein are further metabolically engineered for use in i) Modified expression of endogenous membrane proteins, and/or ii) the modified activity of the endogenous membrane protein, and/or iii) Representation of homologous membrane proteins, and/or iv) expression of heterologous membrane proteins, wherein the membrane protein is involved in the uptake of precursors and/or receptors for the synthesis of any of the 2'FL, 3-FL and DiFL. As explained in the definitions disclosed herein, the terms "precursor" and "acceptor" should be understood. The cell may express one of the membrane proteins involved in the uptake of any type of precursor and/or receptor for the synthesis of any of the 2'FL, 3-FL and DiFL. The cell may also express more than one of the membrane proteins involved in the uptake of at least one of the precursors and/or receptors. The cells can be modified for uptake of more than one precursor and/or receptor for the synthesis of any of the 2'FL, 3-FL and DiFL.

In a specific example, the cells are further genetically modified for use in i) Modified expression of endogenous membrane proteins, and/or ii) the modified activity of the endogenous membrane protein, and/or iii) Representation of homologous membrane proteins, and/or iv) expression of heterologous membrane proteins, wherein the membrane protein is involved in uptake of lactose and/or at least one other receptor 2'FL and/or 3FL for the synthesis of any of the 2'FL, 3-FL and DiFL.

In a more preferred embodiment of the method and/or cell of the present invention, the membrane protein is selected from the list comprising: transporter protein, P-P-bond hydrolysis-driven transporter protein, β-barrel porin, accessory transporter Proteins, putative transport proteins and phosphotransfer driven group translocation proteins. In even more preferred embodiments of the methods and/or cells of the present invention, the transport proteins comprise MFS transport proteins, carbohydrate efflux transport proteins, and chelatin export proteins. In another more preferred embodiment of the method of the present invention, the P-P-bond hydrolysis-driven transporter comprises ABC transporter and chelatin exporter.

In another preferred embodiment of the methods and/or cells of the present invention, the membrane protein provides improved production of any of the 2'FL, 3-FL and DiFL. In alternative and/or additional preferred embodiments of the methods and/or cells of the invention, membrane proteins enable efflux of any of the 2'FL, 3-FL and DiFL.

In a more preferred embodiment of the method and/or cell of the present invention, the cell expresses a membrane protein belonging to the MFS transporter family, such transporter proteins as for example derived from Escherichia coli (UniProt ID P0AEY8), S. The MdfA family of multidrug transporters of species Cronobacter muytjensii (UniProt ID A0A2T7ANQ9), Citrobacter youngae (UniProt ID D4BC23) and Yokenella regensburgei (UniProt ID G9Z5F4) The MdfA polypeptide. In another more preferred embodiment of the method and/or cell of the present invention, the cell expresses a membrane protein belonging to the sugar efflux transporter family, such as for example from Escherichia coli (UniProt ID P31675), SetA polypeptides of the SetA family of species Citrobacter koseri (UniProt ID A0A078LM16) and Klebsiella pneumoniae (UniProt ID A0A0C4MGS7). In another more preferred embodiment of the method and/or cell of the present invention, the cell expresses membrane proteins belonging to the chelatin exporter family, such as, for example, E. coli entS (UniProt ID P24077) and E. coli iceT ( UniProt ID A0A024L207). In another more preferred embodiment of the method and/or cell of the present invention, the cell expresses a membrane protein belonging to the ABC transporter family, such as, for example, oppF from Escherichia coli (UniProt ID P77737), from Lactococcus lactis lmrA (UniProt ID A0A1V0NEL4) from Lactococcus lactis subsp. lactis bv. diacetylactis and Blon_2475 (UniProt ID B7GPD4) from Bifidobacterium longum subsp. infantis.

Optionally, the cell is transformed to comprise at least one nucleic acid sequence encoding a protein selected from the group comprising: lactose transporter, glucose transporter, galactose transporter, fucose transporter, and for nuclear Glycolate-activated sugar transporter, such as for example for GDP-Fuc, wherein the transporter is internalized to precursor-supplemented medium for oligosaccharide synthesis, or for 2'-fucosyllactose , 3-fucosyllactose and/or 2',3-difucosyllactose transport protein of any one.

In another embodiment of the method and/or cell, the cell produces precursors and/or receptors for the synthesis of any of the 2'FL, 3-FL and DiFL. In a preferred embodiment, the cell produces one or more precursors and/or receptors for the synthesis of the 2'FL, 3-FL and DiFL mixture. In a more preferred embodiment, the cell is modified for optimized production of any of the precursors and/or receptors for the synthesis of any of the 2'FL, 3-FL and DiFL. one.

In a specific example, the cell produces lactose for the synthesis of any of the 2'FL, 3-FL and DiFL, preferably the cell further produces the one of the other receptors 2'FL and/or 3FL or at least one of more.

In one embodiment, the present invention provides a method for producing DiFL using cells that fully convert the precursors and/or receptors to DiFL.

In one embodiment, the present invention provides a method for producing a mixture of 2'FL, 3-FL, and DiFL using a cell, wherein the cell fully converts any of the precursors and/or receptors to Any of the 2'FL, 3-FL and DiFL.

In a preferred embodiment of the methods and/or cells of the invention, the cell confers enhanced phage resistance, that is, the cell is more Better to confer higher phage resistance. This enhancement of phage resistance may result from reduced expression of endogenous membrane proteins and/or mutations in genes encoding endogenous membrane proteins. The terms "phage insensitive" or "phage resistant" or "phage resistance" or "phage resistant profile" should be understood to mean and/or growth inhibition of infection and/or killing of less susceptible and preferably less susceptible bacterial strains. As used herein, the term "anti-phage activity" or "resistant to infection by at least one phage" refers to a system that exhibits a functional phage resistance system Increased resistance of bacterial cells to infection by at least one family of bacteriophages, as can be achieved by, for example, bacterial viability, compared to bacterial cells of the same species at the same developmental stage (eg, in culture) that do not exhibit a functional phage resistance system , phage lysogenicity, phage genome replication, and phage genome degradation. Phages can be lytic phages or mild (lysogenic) phages. Membrane proteins involved in phage resistance of cells include OmpA, OmpC, OmpF, OmpT, BtuB, TolC, LamB, FhuA, TonB, FadL, Tsx, FepA, YncD, PhoE and NfrA and their homologues.

In a preferred embodiment of the methods and/or cells of the present invention, the cell confers reduced viscosity, ie the cell preferably confers a reduced viscosity compared to the wild-type precursor cell of the modified cell or metabolically engineered cell lower viscosity. Reduction in cell viscosity can be obtained by biosynthesis of modified cell walls. Cell wall biosynthesis can be modified to include, for example, poly-N-acetyl-glucosamine, enterobacterial common antigen, cellulose, colanic acid, core oligosaccharides, osmoregulatory periplasmic glucans, and glycerol The synthesis of glucosylglycerol, polysaccharides and trehalose is reduced or eliminated.

Another embodiment of the present invention provides a method and a cell wherein 2'FL, 3-FL and DiFL are produced and/or in a fungal, yeast, bacterial, insect, animal, plant or protozoal cell as described herein produced by it. The cell line is selected from a list comprising bacteria, yeast, protozoa or fungi, or refers to plant or animal cells. The latter bacteria preferably belong to the phylum Proteobacteria or Firmicutes or Cyanobacteria or Deinococcus-Thermus. Bacteria belonging to the later phylum Proteobacteria preferably belong to the family Enterobacteriaceae, preferably to the species Escherichia coli. The latter bacteria preferably refer to any strain belonging to the species Escherichia coli, such as, but not limited to, Escherichia coli B, Escherichia coli C, Escherichia coli W, Escherichia coli K12, Escherichia coli Nissle. More specifically, the latter term refers to cultured E. coli strains designated as E. coli K12 strains - which are well adapted to the laboratory environment and which, unlike wild-type strains, have lost their presence in the intestinal tract. the ability to grow. Well-known examples of E. coli K12 strains are K12 wild type, W3110, MG1655, M182, MC1000, MC1060, MC1061, MC4100, JM101, NZN111 and AA200. Therefore, the present invention specifically relates to a mutated and/or transformed E. coli cell or strain as indicated above, wherein the E. coli strain is the K12 strain. The Escherichia coli K12 strain is more preferably Escherichia coli MG1655. Bacteria belonging to the later Firmicutes phylum preferably belong to the Bacilli class, preferably the Lactobacilliales with members such as Lactobacillus lactis , Leuconostoc mesenteroides , or There are members such as Bacillales from the genus Bacillus, such as Bacillus subtilis or B. amyloliquefaciens . The bacteria belonging to the rear of the Actinobacteria phylum preferably belong to the Corynebacteriaceae family with members Corynebacterium glutamicum or C. afermentans , or belong to the family Streptomyces griseus ( Streptomyces griseus ) or the Streptomycetaceae family of S. fradiae . The latter yeast preferably belong to the phylum Ascomycota or Basidiomycota or Deuteromycota or Zygomycetes. The latter yeast preferably belong to the genus Saccharomyces (with members such as, for example, Saccharomyces cerevisiae, S. bayanus , S. boulardii , Zygosaccharomyces ), Pichia ( Pichia) (with members such as eg Pichia pastoris , P. anomala , P. kluyveri ), Komagataella , Hansenula, Kluyveromyces (with, for example, Kluyveromyces lactis , K. marxianus , K. thermotolerans ), Debaromyces , Yarrowia (as for example Yarrowia lipolytica ) or Starmerella (as for example Bumblebee Tamo yeast ( Starmerella bombicola ). The latter yeast is preferably selected from Pichia pastoris , Yarrowia lipolytica , Saccharomyces cerevisiae and Kluyveromyces lactis . The latter fungi preferably belong to the genera Rhizopus, Dictyostelium, Penicillium, Mucor or Aspergillus. Plant cells include cells of flowering and non-flowering plants, as well as algal cells such as Chlamydomonas, Chlorella, and the like. Preferably, the plant is a tobacco, alfalfa, rice, tomato, cotton, rapeseed, soybean, maize or corn plant. The latter animal cells are preferably derived from non-human mammals (eg cattle, buffalo, pigs, sheep, mice, rats), birds (eg chickens, ducks, ostriches, turkeys, pheasants), fish (eg swordfish, salmon, tuna, sea bass, trout, catfish), invertebrates (e.g. lobster, crab, shrimp, clams, oysters, mussels, sea urchins), reptiles (e.g. snakes, crocodiles, turtles), amphibians ( eg frogs) or insects (eg flies, nematodes) or are genetically modified cell lines derived from human cells devoid of embryonic stem cells. Both human and non-human mammalian cells are preferably selected from a list comprising: epithelial cells such as eg mammary epithelial cells, embryonic kidney cells (eg HEK293 or HEK 293T cells), fibroblasts, COS cells, Chinese hamster ovary ( Chinese hamster ovary; CHO) cells, murine myeloma cells such as eg N20, SP2/0 or YB2/0 cells, NIH-3T3 cells, non-mammary adult stem cells or derivatives thereof, such as described in WO21067641. The latter insect cells are preferably derived from: Spodoptera frugiperda such as eg Sf9 or Sf21 cells, Bombyx mori , Mamestra brassicae , Trichoplusia ni such as eg BTI- TN-5B1-4 cells or Drosophila melanogaster such as Drosophila S2 cells. The latter protozoal cells are preferably Leishmania tarentolae cells.

In a preferred embodiment of the methods and/or cells of the invention, the cells are viable Gram-negative bacteria comprising reduced or eliminated synthetic poly-N- Acetyl-glucosamine (poly-N-acetyl-glucosamine; PNAG), Enterobacterial Common Antigen (ECA), cellulose, kolaric acid, core oligosaccharide, osmoregulatory periplasmic glucan ( Osmoregulated Periplasmic Glucan; OPG), glycerol glucosides, glycans and/or trehalose.

In a more preferred embodiment of the method and/or cell, the reduction or elimination of synthetic poly-N-acetyl-glucosamine (PNAG), enterobacterial common antigen (ECA), cellulose, kolac acid, Core oligosaccharides, osmoregulatory periplasmic glucans (OPG), glycerol glucosides, glycans and/or trehalose are produced by participating in the poly-N-acetyl-glucosamine (PNAG), enterobacterial common antigen Synthesis of any of (ECA), cellulose, kolac acid, core oligosaccharides, osmoregulatory periplasmic glucans (OPGs), glycerol glucosides, polysaccharides, and/or trehalose Mutations of glycosyltransferases provide for deletion or lower performance of any of the glycosyltransferases. Compared with unmodified precursor cells, these glycosyltransferases comprise a glycosyltransferase gene encoding a poly-N-acetyl-D-glucosamine synthase subunit, UDP-N-acetylglucosamine -Undecyl isopentenyl-phosphate N-acetylglucosamine phosphotransferase, Fuc4NAc (4-acetamido-4,6-dideoxy-D-galactose) transferase, UDP-N-ethyl Acyl-D-mannuronosyltransferase, glycosyltransferase gene encoding the catalytic subunit of cellulose synthase, cellulose biosynthesis protein, colaric acid biosynthesis glucuronyltransferase, colaric acid biosynthesis Galactosyltransferase, Kolaic acid biosynthetic fucosyltransferase, UDP-glucose:undecyl isopentenyl-phosphoglucosyl-1-phosphotransferase, putative Kolaic acid biosynthetic glycosyltransferase, UDP - Glucuronic acid: LPS(HepIII) glycosyltransferase, ADP-heptose-LPS heptosyltransferase 2, ADP-heptose:LPS heptosyltransferase 1, putative ADP-heptose:LPS heptose Syltransferase 4, lipopolysaccharide core biosynthesis protein, UDP-glucose:(glucosyl)LPS α-1,2-glucosyltransferase, UDP-D-glucose:(glucosyl)LPS α-1,3-glucose Syltransferase, UDP-D-galactose: (glucosyl) lipopolysaccharide-1,6-D-galactosyltransferase, lipopolysaccharide glucosyltransferase I, lipopolysaccharide core heptosyltransferase 3,β- 1,6-Galactosefuranosyltransferase, Undecyl isopentenyl-phosphate 4-deoxy-4-carbamoylamino-L-arabinosyltransferase, Lipid IVA 4-amino-4-deoxy -L-arabinosyltransferase, bacterioterpene glucosyltransferase, putative family 2 glycosyltransferase, osmoregulatory periplasmic glucan (OPG) biosynthesis protein G, OPG biosynthesis protein H, glucosylglycerol Ester phosphorylase, liver sugar synthase, 1,4-α-glucan branching enzyme, 4-α-glucanotransferase and trehalose-6-phosphate synthase. In an illustrative embodiment, the cell is mutated in any one or more of the glycosyltransferases comprising pgaC, pgaD, rfe, rffT, rffM, bcsA, bcsB, bcsC , wcaA, wcaC, wcaE, wcaI, wcaJ, wcaL, waaH, waaF, waaC, waaU, waaZ, waaJ, waaO, waaB, waaS, waaG, waaQ, wbbl, arnC, arnT, yfdH, wbbK, opgG, opgH, ycjM , glgA, glgB, malQ, otsA, and yaiP, wherein the mutation provides deletion or lower expression of any of the glycosyltransferases.

In alternative and/or additional preferred embodiments of the method and/or cell, the reduction or elimination of poly-N-acetyl-glucosamine (PNAG) synthesis by overexpression of genes encoding carbon storage regulators, Na+ Deletion of the gene encoding the /H+ antiporter regulator and/or deletion of the gene encoding the sensor histidine kinase is provided.

Microorganisms or cells as used herein can be grown on monosaccharides, disaccharides, oligosaccharides, polysaccharides, polyols, glycerol; complex media including molasses, corn infusion, meringue, pancreas, yeast extract, or mixtures thereof (eg, mixed Feedstocks, preferably mixed monosaccharide feedstocks such as, for example, hydrolyzed sucrose) are grown on as the main carbon source. The term "complex medium" means a medium in which the exact composition is not defined. The term primarily means the most important carbon source for the biological product of interest, biomass formation, carbon dioxide and/or by-product formation (such as acids and/or alcohols, such as acetate, lactate and/or ethanol), i.e. all required 20%, 30%, 40%, 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 98%, 99% of the carbon is derived from the carbon sources indicated above. In one embodiment of the present invention, the carbon source is the sole carbon source for the organism, ie 100% of all required carbon is derived from the carbon sources indicated above. Common major carbon sources include (but are not limited to) glucose, glycerol, fructose, maltose, lactose, arabinose, malto-oligosaccharide, maltotriose, sorbitol, xylose, rhamnose, sucrose, galactose, mannose Sugar, methanol, ethanol, trehalose, starch, cellulose, hemicellulose, molasses, corn infusion, high fructose syrup, acetate, citrate, lactate and pyruvate. The term complex medium means a medium in which the exact composition is not defined. Examples are molasses, corn steep liquor, meringue, pancreas or yeast extract.

In another preferred embodiment, the microorganisms or cells described herein use split metabolism with production pathways and biomass pathways as described in WO2012/007481, which is incorporated herein by reference. The organism may be genetically modified, for example, by altering a gene selected from the group consisting of a phosphoglucose isomerase gene, a phosphofructokinase gene, a fructose-6-phosphate aldolase gene, a fructose isomerase gene and/or a fructose:PEP phosphotransferase gene for gene to accumulate fructose-6-phosphate.

According to another embodiment of the method and/or cell of the present invention, the conditions that allow for the production of the DiFL or the mixture of 2'FL, 3-FL and DiFL comprise the use of a mixture comprising at least one for the production of the DiFL or the 2'FL, 3 - FL and DiFL mixture for precursor and/or acceptor medium. Preferably, the medium contains at least one precursor selected from the group consisting of lactose, galactose, and fucose.

According to an alternative and/or additional embodiment of the method of the present invention, the conditions permitting the production of the DiFL or the mixture of 2'FL, 3-FL and DiFL comprise adding at least one precursor and/or acceptor feed to the culture medium to The DiFL or the mixture of 2'FL, 3-FL and DiFL is produced.

According to an alternative embodiment of the method of the present invention, the conditions allowing the production of the DiFL or the mixture of 2'FL, 3-FL and DiFL comprise the use of a medium for culturing a culture medium for the production of DiFL or the mixture of 2'FL, 3-FL and DiFL Cells of the invention, wherein the medium lacks any precursor and/or receptor for the production of the DiFL or the mixture of 2'FL, 3-FL and DiFL, and is added to cells having at least one precursor and/or receptor for the production of the DiFL or the 2'FL The 'FL, 3-FL and DiFL mixtures were combined with another supplement in the medium of the precursor and/or acceptor feed.

In a preferred embodiment, the method for producing DiFL as described herein or the mixture of 2'FL, 3-FL and DiFL comprises at least one of the following steps: i) using a method comprising at least one precursor and/or or acceptor medium; ii) adding at least one precursor and/or acceptor feed to the medium in a reactor with a total reactor volume in the range of 250 mL (milliliters) to 10.000 ^m3 (cubic meters) , preferably in a continuous manner, and preferably such that the final volume of the medium is no more than three times, preferably no more than twice, more preferably less than two times the volume of the medium before the addition of the precursor and/or acceptor feed iii) adding at least one precursor and/or acceptor feed to the culture medium in a reactor, wherein the total reactor volume is in the range of 250 mL (milliliters) to 10.000 ^m3 (cubic meters), preferably with is added in a continuous manner, and preferably such that the final volume of the medium is no more than three times, preferably no more than two times, more preferably less than two times the volume of the medium before the addition of the precursor and/or acceptor feed, and wherein more Preferably, the pH of the precursor and/or acceptor feed is set between 3 and 7, and wherein preferably the temperature of the precursor and/or acceptor feed is maintained between 20°C and 80°C iv) at least one precursor and/or acceptor feed is added to the medium in a continuous manner by means of a feed solution over a time course of 1 day, 2 days, 3 days, 4 days, 5 days; v) over a period of 1 day , 2-day, 3-day, 4-day, 5-day time course to add at least one precursor and/or acceptor feed to the medium by means of a feed solution in a continuous manner, and wherein preferably, the feed solution is The pH is set between 3 and 7, and wherein preferably the temperature of the feed solution is maintained between 20°C and 80°C; the method yields a concentration of at least 50 g/L, preferably at least 75 g/L in the final culture g/L, more preferably at least 90 g/L, more preferably at least 100 g/L, more preferably at least 125 g/L, more preferably at least 150 g/L, more preferably at least 175 g/L, more preferably at least 200 g/L DiFL of L or any of the 2'FL, 3-FL and DiFL in the mixture of 2'FL, 3-FL and DiFL.

In another and/or additional preferred embodiment, the method for producing DiFL as described herein or the mixture of 2'FL, 3-FL and DiFL comprises at least one of the following steps: i) using a method comprising each Liter initial reactor volume of at least 50, more preferably at least 75, more preferably at least 100, more preferably at least 120, more preferably at least 150 grams of lactose culture medium, wherein the reactor volume is between 250 mL and 10.000 m ³ (cubic meters) ii) Add at least one precursor and/or acceptor to the culture medium in one pulse or discontinuously (pulsed) with a total reactor volume ranging from 250 mL (milliliter) to 10.000 ^m3 (cubic meter). ), preferably such that the final volume of the medium does not exceed three times, preferably not more than twice, more preferably less than twice the volume of the medium before adding the precursor and/or acceptor feed pulse; iii) with Add at least one precursor and/or acceptor feed to the culture medium in a pulsed or discontinuous (pulsed) manner in a reactor, wherein the total reactor volume is between 250 mL (milliliter) and 10.000 m ³ (cubic g). feet), preferably such that the final volume of the medium is no more than three times, preferably no more than two times, more preferably less than two times the volume of the medium before the addition of the precursor and/or acceptor feed pulse, and wherein Preferably, the pH of the precursor and/or acceptor feed pulse is set between 3 and 7, and wherein preferably the temperature of the precursor and/or acceptor feed pulse is maintained between 20°C and 80°C. °C; iv) over a time course of 5 minutes, 10 minutes, 30 minutes, 1 hour, 2 hours, 4 hours, 10 hours, 12 hours, 1 day, 2 days, 3 days, 4 days, 5 days by means of Feed solution adding at least one precursor and/or acceptor feed to the culture medium in a discontinuous (pulsed) manner; v) over 5 minutes, 10 minutes, 30 minutes, 1 hour, 2 hours, 4 hours, 10 Add at least one precursor and/or acceptor feed to the medium in a discontinuous (pulsed) manner by means of a feed solution over a time course of hours, 12 hours, 1 day, 2 days, 3 days, 4 days, 5 days , and wherein preferably the pH of the feed solution is set between 3 and 7, and wherein preferably the temperature of the feed solution is maintained between 20°C and 80°C; the method produces in the final culture The concentration is at least 50 g/L, preferably at least 75 g/L, more preferably at least 90 g/L, more preferably at least 100 g/L, more preferably at least 125 g/L, more preferably at least 150 g/L, more preferably At least 175 g/L, more preferably at least 200 g/L of DiFL or any of the 2'FL, 3-FL and DiFL in the mixture of 2'FL, 3-FL and DiFL.

In another more preferred embodiment, the method for producing DiFL as described herein or the mixture of 2'FL, 3-FL and DiFL comprises at least one of the following steps: i) using an initial reactor comprising per liter a medium having a volume of at least 50, more preferably at least 75, more preferably at least 100, more preferably at least 120, more preferably at least 150 grams of lactose, wherein the reactor volume is in the range of 250 mL to 10.000 m ³ (cubic meters); ii ) adding to the medium a lactose feed comprising at least 50, more preferably at least 75, more preferably at least 100, more preferably at least 120, more preferably at least 150 grams of lactose per liter of initial reactor volume, wherein the total reactor volume is at 250 mL (milliliters) to 10.000 ^m3 (cubic meters), preferably in a continuous manner, and preferably such that the final volume of the medium does not exceed three times the volume of the medium before the addition of the lactose feed, preferably not more than twice, more preferably less than 2 times; iii) adding to the medium lactose comprising at least 50, more preferably at least 75, more preferably at least 100, more preferably at least 120, more preferably at least 150 grams of lactose per liter of initial reactor volume feed, wherein the reactor volume is in the range of 250 mL to 10.000 ^m3 (cubic meters), preferably in a continuous manner, and preferably such that the final volume of the medium does not exceed the volume of the medium prior to addition of the lactose feed three times, preferably no more than twice, more preferably less than 2 times, and wherein preferably the pH of the lactose feed is set between 3 and 7, and wherein preferably the temperature of the lactose feed is maintained between 20°C and 80°C; iv) over a time course of 1 day, 2 days, 3 days, 4 days, 5 days, to the medium by means of the feed solution a lactose feed was added in a continuous manner; v) over a period of 1 day , 2 days, 3 days, 4 days, 5 days time course by means of a feed solution to add lactose feed to the medium in a continuous manner, and wherein the concentration of the lactose feed solution is 50 g/L, preferably 75 g /L, better 100 g/L, better 125 g/L, better 150 g/L, better 175 g/L, better 200 g/L, better 225 g/L, better 250 g/ L, better 275 g/L, better 300 g/L, better 325 g/L, better 350 g/L, better 375 g/L, better 400 g/L, better 450 g/L , better 500 g/L, even better 550 g/L, best 600 g/L; and wherein preferably, the pH of the feed solution is set between 3 and 7, and wherein preferably, the The temperature of the feed solution is maintained between 20°C and 80°C; the method produces a concentration in the final culture of at least 50 g/L, preferably at least 75 g/L, more preferably at least 90 g/L, more preferably at least 100 g/L g/L, preferably at least 12 5 g/L, more preferably at least 150 g/L, more preferably at least 175 g/L, more preferably at least 200 g/L of DiFL or the 2'FL, 3 of the mixture of 2'FL, 3-FL and DiFL - Any of FL and DiFL.

Preferably, the lactose is fed by starting the culture at a concentration of at least 5 mM, preferably at a concentration of 30, 40, 50, 60, 70, 80, 90, 100, 150 mM, more preferably at > This was achieved by adding lactose at a concentration of 300 mM.

In another embodiment, lactose feeding is achieved by adding lactose to the medium at a concentration such that a lactose concentration of at least 5 mM, preferably 10 mM or 30 mM is obtained throughout the production phase of the culture.

In another embodiment of the methods described herein, the host cells are cultured for at least about 60, 80, 100, or about 120 hours or in a continuous manner.

In a preferred embodiment, the carbon source, preferably sucrose, is provided in the medium for 3 or more days, preferably up to 7 days; and/or the medium is provided in a continuous manner per liter of initial culture volume at least 100. Advantageously at least 105, more advantageously at least 110, even more advantageously at least 120 grams of sucrose, such that the final volume of the medium is no more than three times, advantageously no more than twice, and more advantageously less than two times the volume of the medium before culturing.

Preferably, when carrying out the method as described herein, the first stage of exponential cell growth is carried out by adding a carbon source, preferably glucose or sucrose, before adding the precursor to the culture in the second stage provided in the medium.

In another preferred embodiment of the method of the present invention, the first stage of exponential cell growth is provided by adding a carbon-based substrate, preferably glucose or sucrose to the medium containing the precursor, followed by a A carbon-based substrate, preferably glucose or sucrose, is added to the second stage in the culture.

In another preferred embodiment of the method of the invention, the first stage of exponential cell growth is provided by adding a carbon-based substrate, preferably glucose or sucrose to the medium comprising the precursor, followed by A carbon-based substrate, preferably glucose or sucrose, and precursors are added to the second stage in the culture.

In an alternative preferred embodiment, in the method as described herein, the precursor has been added with the carbon-based substrate in the first stage of exponential growth.

According to the present invention, the method as described herein preferably comprises isolating at least one of the 2'FL, 3-FL and DiFL from the culture or reaction mixture, more preferably isolating the 2' from the culture or reaction mixture All steps in FL, 3-FL and DiFL.

The term "separating from said cultivation or reaction mixture" means capturing, collecting or obtaining the 2'FL, 3 - any of FL and DiFL, preferably all of the 2'FL, 3-FL and DiFL.

Any of the 2'FL, 3-FL and DiFL can be isolated from the aqueous medium in which the cells are grown in a conventional manner. In cases where the 2'FL, 3-FL and/or DiFL are still present in the cells producing the mixture of 2'FL, 3-FL and DiFL, the release or extraction of the 2'FL, 3-FL from the cells can be used and/or DiFL in conventional manner, such as using high pH, heat shock, sonication, French press, homogenization, enzymatic hydrolysis, chemical hydrolysis, solvent hydrolysis, detergents, hydrolysis... to destroy the cells. The culture medium and/or cell extract and/or enzyme mixture together and respectively can then be further used to isolate the 2'FL, 3-FL and/or DiFL. This preferably involves clarification of the mixture containing the 2'FL, 3-FL and DiFL to remove suspended particles and contaminants, especially cells, cellular components, insoluble metabolites and debris produced by culturing genetically modified cells. In this step, the mixture containing the 2'FL, 3-FL and DiFL can be clarified in a known manner. Preferably, the mixture containing the 2'FL, 3-FL and DiFL is clarified by centrifugation, flocculation, decantation and/or filtration. Another step of separating the 2'FL, 3-FL and/or DiFL from the mixture containing the 2'FL, 3-FL and DiFL preferably involves removal from the mixture containing the 2'FL, 3-FL and DiFL Substantially all proteins, as well as peptides, amino acids, RNA and DNA and any endotoxins and glycolipids that could interfere with subsequent separation steps, are preferably after they have been clarified. In this step, protein and related impurities can be removed from the mixture containing the 2'FL, 3-FL and DiFL in a conventional manner. Preferably, proteins, salts, by-products, dyes, endotoxins and other related impurities are filtered from the mixture containing the 2'FL, 3-FL and DiFL by ultrafiltration, nanofiltration, biphasic partition, reverse osmosis, microfiltration Filtration, activated carbon or carbon treatment, tangential flow high performance filtration, treatment with non-ionic surfactants, enzymatic digestion, tangential flow ultrafiltration, electrophoresis (e.g. using slabs - polyacrylamide or dodecyl sulfate) sodium-polyacrylamide gel electrophoresis (PAGE)), affinity chromatography (using affinity ligands, including, for example, DEAE-agarose, poly-L-lysine, and polymyxin- B. Endotoxin-selective adsorbent matrix), ion exchange chromatography (such as (but not limited to) cation exchange, anion exchange, mixed bed ion exchange, ligand attachment from the inside out), hydrophobic interaction chromatography and/or gel filtration (ie size exclusion chromatography), specifically by chromatography, more specifically by ion exchange chromatography or hydrophobic interaction chromatography or ligand exchange chromatography remove. In addition to size exclusion chromatography, the 2'FL, 3-FL and DiFL are retained in mixtures containing the 2'FL, 3-FL and DiFL by chromatographic media or membranes of choice to retain proteins and related impurities middle.

In another preferred embodiment, the methods as described herein also provide further purification of any one or more of 2'FL, 3-FL and/or DiFL from a mixture of 2'FL, 3-FL and DiFL. Further purification of the 2'FL, 3-FL and/or DiFL can e.g. by using (activated) charcoal or carbon, nanofiltration, ultrafiltration or ion exchange to remove any residual DNA, protein, LPS, endotoxin or other impurities to achieve. Alcohols such as ethanol and hydroalcoholic mixtures can also be used. Another purification step is achieved by crystallization, evaporation or precipitation of the product. Another purification step is drying the resulting oligosaccharides, e.g. spray drying, freeze drying, spray freeze drying, freeze spray drying, band dry, belt dry, vacuum belt drying Drying, vacuum belt drying, tumble drying, tumble drying, vacuum tumble drying or vacuum tumble drying.

In a specific example, the resulting DiFL is purified from the mixture of 2'FL, 3-FL, and DiFL to the purity of the 2'FL, 3-FL produced in the mixture of 2'FL, 3-FL, and DiFL. The total of FL and DiFL was measured to be at least 80%.

In an illustrative embodiment, the isolation and purification of at least one, preferably all of the resulting 2'FL, 3-FL and DiFL is performed in a process comprising the following steps in any order: a) Contact the culture or its clarified form or enzymatic reaction mixture with a nanofiltration membrane with a molecular weight cut-off (MWCO) of 600-3500 Da, ensuring retention of the 2'FL, 3-FL produced and/or DiFL and pass at least a portion of proteins, salts, by-products, dyes and other related impurities, b) Using the membrane, the retentate from step a) is subjected to a diafiltration process with an aqueous solution of inorganic electrolyte, followed by diafiltration with pure water as appropriate to remove excess electrolyte, c) and collecting the oligosaccharide-enriched retentate(s) in salt form from the cations of the electrolyte. d) Preferably, the retentate is dried by any one or more drying steps selected from the list comprising: spray drying, freeze drying, spray freeze drying, freeze spray drying, strip drying, belt drying, Vacuum belt drying, vacuum belt drying, roller drying, roller drying, vacuum roller drying and vacuum roller drying.

In an alternative illustrative embodiment, the isolation and purification of at least one, preferably all of the resulting 2'FL, 3-FL and DiFL is performed in a process comprising the following steps, in any order: The culture or its clarified form or reaction mixture is subjected to two membrane filtration steps using different membranes, wherein one membrane has a molecular weight cut-off between about 300 and about 500 Daltons and the other has a molecular weight cut-off between about 600 and about 800 between Daltons.

In an alternative illustrative embodiment, the isolation and purification of at least one of the 2'FL, 3-FL and DiFL produced is performed in a process comprising the following steps in any order, including the following steps : Treatment of the culture or its clarified form or reaction mixture with a strong cation exchange resin in the H+ form and a weak anion exchange resin in the free base form.

In an alternative illustrative embodiment, the isolation and purification of at least one of the produced 2'FL, 3-FL and DiFL is performed in the following manner. Cultures containing produced 2'FL, 3-FL and DiFL, biomass, media components and contaminants were applied to the following purification steps: i) separation of biomass from the culture, ii) cation exchanger treatment for removal of positively charged species, iii) anion exchanger treatment for removal of negatively charged species, iv) performing a nanofiltration step and/or an electrodialysis step, Provided therein is a purified solution comprising the desired oligosaccharide produced from a mixture of 2'FL, 3-FL and/or DiFL with a purity greater than or equal to 80%. Optionally, the purified solution is dried by any one or more drying steps selected from the list comprising: spray drying, lyophilization, spray freeze drying, freeze spray drying, ribbon drying, ribbon drying, vacuum ribbon Drying, vacuum belt drying, drum drying, drum drying, vacuum drum drying and vacuum drum drying.

In a specific example, the isolation and purification of all of the 2'FL, 3-FL and DiFL produced is performed in the following manner. Cultures containing produced 2'FL, 3-FL and DiFL, biomass, media components and contaminants were applied to the following purification steps: i) separation of biomass from the culture, ii) cation exchanger treatment for removal of positively charged species, iii) anion exchanger treatment for removal of negatively charged species, iv) performing a nanofiltration step and/or an electrodialysis step, Provided therein is a purified solution comprising the resulting mixture of 2'FL, 3-FL and/or DiFL with a combined purity greater than or equal to 80%. Optionally, the purified solution is dried by any one or more drying steps selected from the list comprising: spray drying, lyophilization, spray freeze drying, freeze spray drying, ribbon drying, ribbon drying, vacuum ribbon Drying, vacuum belt drying.

In a preferred embodiment, the present invention provides for the production of DiFL having a purity of at least 80% as measured by the total amount of 2'FL, 3-FL and DiFL produced in the mixture. As used herein, a DiFL purity of at least 80% is understood to mean 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 95.5, 96, 96.5, 97, 97.5, 98, 98.5, 99 or 99.5% pure.

In an alternative illustrative embodiment, the isolation of at least one of the 2'FL, 3-FL, and DiFL produced can be performed in a process comprising the following steps, in any order: Enzymatic treatment; removal of biomass from culture; ultrafiltration; nanofiltration; and performing column chromatography steps. Preferably, such column chromatography is a single column or multiple columns. More preferably, the column chromatography step is simulated moving bed chromatography. Such simulated moving bed chromatography preferably comprises i) at least 4 columns, at least one of which comprises a weak or strong cation exchange resin; and/or ii) four zones I, II, III and IV with different flow rates ; and/or iii) a dissolving agent comprising water; and/or iv) an operating temperature of 15 to 60 degrees Celsius. Preferably, the process further comprises a drying step selected from the list comprising: spray drying, freeze drying, spray freeze drying, freeze spray drying, strip drying, belt drying, vacuum strip drying, vacuum strip drying Type drying, tumble drying, tumble drying, vacuum tumble drying and vacuum tumble drying.

In one embodiment, the present invention provides the DiFL produced, which is dried to powder by any one or more drying steps selected from the list comprising: spray drying, lyophilization, spray freeze drying, freeze spray drying, Strip drying, belt drying, vacuum strip drying, vacuum belt drying, drum drying, drum drying, vacuum drum drying and vacuum drum drying, wherein the dry powder contains < 15 wt.% water, preferably < 10 wt .% water, better < 7 wt. % water, best < 5 wt. % water.

In a specific embodiment, the present invention provides 2'FL, 3-FL and/or DiFL or mixtures of 2'FL, 3-FL and DiFL produced by any one selected from a list comprising: or Multiple drying steps to powder: spray drying, freeze drying, spray freeze drying, freeze spray drying, strip drying, belt drying, vacuum strip drying, vacuum belt drying, drum drying, roller drying, vacuum drum Drying and vacuum roller drying wherein the dry powder contains < 15 wt. % water, preferably < 10 wt. % water, more preferably < 7 wt. % water, most preferably < 5 wt. % water.

In a third aspect, the present invention provides the use of metabolically engineered cells as described herein for the production of DiFL. In another aspect, the present invention provides the use of a method for producing DiFL.

In a specific example, the present invention provides the use of metabolically engineered cells as described herein for the production of 2'FL, 3-FL and DiFL mixtures. In another aspect, the present invention provides the use of a method for producing a mixture of 2'FL, 3FL and DiFL.

In a specific example, the present invention provides the use of metabolically engineered cells as described herein for the production of 2'FL and DiFL mixtures. In another aspect, the present invention provides the use of a method for producing a mixture of 2'FL and DiFL.

In a specific example, the present invention provides the use of metabolically engineered cells as described herein for the production of 3-FL and DiFL mixtures. In another aspect, the present invention provides the use of a method for producing a mixture of 3FL and DiFL.

To identify saccharides in mixtures comprising 2'FL, 3-FL and/or DiFL produced as described herein, monomer building blocks (e.g. monosaccharide or glycan unit composition), side chain changes Rotational configuration, presence and position of substituents, degree of polymerization/molecular weight, and linkage pattern can be identified by standard methods known in the art, such as, for example, methylation analysis, reductive cleavage, hydrolysis, Gas chromatography-mass spectrometry (GC-MS), matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS), electrospray Electrospray ionization-mass spectrometry (ESI-MS), High-Performance Liquid chromatography (HPLC) with UV or refractive index detection, and high-performance anion exchange with pulsed amperometric detection Chromatography (High-Performance Anion-Exchange chromatography with Pulsed Amperometric Detection; HPAEC-PAD), capillary electrophoresis (CE), infrared (IR)/Raman (Raman) spectroscopy and nuclear magnetic resonance; NMR) spectroscopy. Crystal structures can be solved using, for example, solid-state NMR, Fourier transform infrared (FT-IR) spectroscopy, and wide-angle X-ray scattering (WAXS). The degree of polymerization (DP), DP distribution and polydispersity can be determined, for example, by viscometry and SEC (SEC-HPLC, high performance size exclusion chromatography). To identify the monomeric components of sugars, methods such as, for example, acid-catalyzed hydrolysis, high performance liquid chromatography (HPLC) or gas-liquid chromatography (GLC) (in conversion after aldol acetate). To determine the glycosidic bond, the sugars were methylated in DMSO with methyl iodide and a strong base, hydrolyzed to achieve reduction of partially methylated sugar alcohols, acetylated of methylated aldol acetates, and acetylated by GLC. /MS (gas-liquid chromatography combined with mass spectrometry) for analysis. To sequence oligosaccharides, partial depolymerization is performed using acids or enzymes to determine structure. To identify the mutator configuration, oligosaccharides are subjected to enzymatic analysis, for example, by contacting them with enzymes specific for a particular type of linkage, such as β-galactosidase or α-glucosidase, and NMR. Can be used to analyze products. Products containing oligosaccharide mixture or DiFL

In some embodiments, the oligosaccharide mixture or DiFL produced as described herein is incorporated into a food product (eg, human food or feed), dietary supplement, pharmaceutical ingredient, cosmetic ingredient, or pharmaceutical product. In some embodiments, the oligosaccharide mixture or DiFL is mixed with one or more ingredients suitable for use in food, food, dietary supplements, pharmaceutical ingredients, cosmetic ingredients, or pharmaceuticals.

In some embodiments, the dietary supplement includes at least one probiotic ingredient and/or at least one probiotic ingredient.

"Prebiotics" are substances that promote the growth of microorganisms that are beneficial to the host, especially those in the gastrointestinal tract. In some embodiments, dietary supplements provide a variety of probiotic cobiotics, including oligosaccharide mixtures produced and/or purified by the processes disclosed in this specification, to promote the growth of one or more beneficial microorganisms. Examples of probiotic components of dietary supplements include other probiotic molecules (such as HMO) and plant polysaccharides (such as inulin, pectin, b-glucans and xylo-oligosaccharides). "Probiotic" products typically contain live microorganisms that replace or add to the microbiota of the gastrointestinal tract for the benefit of the recipient. Examples of such microorganisms include Lactobacillus species (eg, L. acidophilus and L. bulgaricus ), Bifidobacterium species (eg, B. animalis ), B. longum and B. infantis (eg Bi-26) and Saccharomyces boulardii . In some embodiments, mixtures of oligosaccharides produced and/or purified by the procedures of this specification are administered orally in combination with such microorganisms.

Examples of other ingredients of dietary supplements include disaccharides (such as lactose), monosaccharides (such as glucose and galactose), thickeners (such as acacia), acidity regulators (such as trisodium citrate), water, skim milk and flavorings.

In some embodiments, the oligosaccharide mixture or DiFL is incorporated into human infant food (eg, infant formula). Infant formula is generally a manufactured food intended for feeding infants as a complete or partial substitute for human breast milk. In some embodiments, infant formula is sold in powder form and prepared by mixing with water for bottle feeding or cup feeding to infants. The composition of infant formula is typically designed to approximately mimic human breast milk. In some embodiments, mixtures of oligosaccharides produced and/or purified by the methods of this specification are included in infant formulas to provide nutritional benefits similar to those provided by oligosaccharides in human breast milk. In some embodiments, the oligosaccharide mixture or DiFL is mixed with one or more ingredients of the infant formula. Examples of infant formula ingredients include skim milk, carbohydrate sources such as lactose, protein sources such as whey protein concentrate and casein, fat sources such as vegetable oils such as palm oil, high oleic safflower oil, canola oil, coconut oil and/or sunflower oil; and fish oil), vitamins (such as vitamins A, Bb, Bi2, C and D), minerals (such as potassium citrate, calcium citrate, magnesium chloride, sodium chloride, citric acid) sodium and calcium phosphate) and possibly human milk oligosaccharide (HMO). Such HMOs may include, for example, DiFL, lacto-N-triose II, LNT, LNnT, lacto-N-fucopentaose I, lacto-N-neofucopentose, lacto-N-fucopentose II, Lacto-N-fucopentaose III, Lacto-N-fucopentaose V, Lacto-N-neofucopentose V, Lacto-N-difucohexaose I, Lacto-N-difucohexaose Sugar II, 6'-galactosylose, 3'-galactosyllose, lacto-N-hexaose and lacto-N-neohexaose.

In some embodiments, the one or more infant formula ingredients comprise skim milk, a carbohydrate source, a protein source, a fat source, and/or vitamins and minerals.

In some embodiments, the one or more infant formula ingredients comprise lactose, whey protein concentrate, and/or high oleic safflower oil.

In some embodiments, the concentration of the oligosaccharide mixture or DiFL in the infant formula is about the same concentration as the concentration of oligosaccharides typically present in human breast milk. In some embodiments, the concentration of each single oligosaccharide in the mixture of oligosaccharides in the infant formula is about the same concentration as the concentration of oligosaccharides typically present in human breast milk.

In some embodiments, the oligosaccharide mixture or DiFL is incorporated into a feed formulation, wherein the feed is selected from a list comprising pet food, animal milk replacer, veterinary product, post-weaning feed, or nursery feed.

Each specific example disclosed in the context of one aspect of the invention is also disclosed in the context of all other aspects of the invention, unless expressly stated otherwise.

Unless otherwise defined, all technical and scientific terms used herein generally have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. In general, the nomenclature used herein and the experimental procedures in cell culture, molecular genetics, organic chemistry and nucleic acid chemistry and hybridization described above and below are those well known and commonly used in the art, in cell culture Experimental procedures, molecular genetics, organic chemistry and nucleic acid chemistry and hybridization. Nucleic acid and peptide synthesis is performed using standard techniques. Generally, purification steps are carried out according to the manufacturer's instructions.

Other advantages arise from specific examples and embodiments. It goes without saying that the features mentioned above and those still to be explained below can be used not only in the respectively specified combination but also in other combinations or independently without departing from the scope of the present invention.

The present invention relates to the following specific embodiments: 1. A method for the production of 2',3-difucosyllactose (DiFL), the method comprising the steps of: i) providing at least two fucosyltransferases , preferably at least one α-1,2-fucosyltransferase and at least one α-1,3-fucosyltransferase, and a GDP-fucose donor, wherein these fucosyltransferases Enzymes capable of transferring fucose residues from the GDP-fucose donor to one or more acceptors, and, ii) linking the fucosyltransferases and the GDP-fucose donor with the containing acceptor A mixture of lactose and possibly one or more other acceptors is contacted under conditions in which the fucosyltransferases catalyze the transfer of fucose residues from the GDP-fucose donor to the acceptor(s) by This produces DiFL and possibly 2'FL and/or 3FL, iii) preferably, at least one of the 2'FL, 3-FL and DiFL is separated, more preferably the 2'FL, 3-FL are separated and all of DiFL. 2. The method of embodiment 1, wherein the DiFL is present in the mixture at a purity of at least 80% as measured by the total amount of 2'FL, 3-FL and DiFL produced in the reaction mixture. 3. The method of any one of embodiments 1 or 2, wherein the fucosyltransferases and the GDP-fucose are provided in a cell-free system. 4. The method of any one of embodiments 1 or 2, wherein the fucosyltransferases and the GDP-fucose are engineered by cells, preferably single cells, preferably metabolically for use in Single cells producing DiFL or a mixture of 2'FL, 3-FL and DiFL were provided. 5. The method of any one of embodiments 1 to 4, wherein the other acceptor(s) is selected from a list comprising 2'FL and 3-FL. 6. The method of any one of Embodiments 1, 2, 4 or 5, comprising the steps of: a. providing a cell, preferably a single cell, wherein the cell expresses at least two fucosyltransferases, more Preferably at least one alpha-1,2-fucosyltransferase and at least one alpha-1,3-fucosyltransferase capable of converting fucose residues from GDP- The alginose donor is transferred to one or more of the acceptor lactose and possibly the other acceptor(s), and the cell is capable of synthesizing GDP-fucose, wherein the GDP-fucose is used for the rock a donor of fucosyltransferases, and b. culturing the cells under conditions that allow expression of the fucosyltransferases and synthesis of the GDP-fucose, c. preferably, isolating the 2 from the culture At least one of 'FL, 3-FL or DiFL, more preferably, all of the 2'FL, 3-FL and DiFL are isolated from the culture. 7. The method of any one of specific examples 4 to 6, wherein the cell expresses: a. at least one α-1,2-fucosyltransferase, the fucosyltransferase (i) comprising as SEQ The polypeptide sequence of any one of ID NOs: 16 to 110, or (ii) is a functional homologue, variant or derivative of any one of SEQ ID NOs 16 to 110, the functional homologue, variant or derivatives having at least 80% overall sequence identity to the full length of any of the alpha-1,2-fucosyltransferase polypeptides having SEQ ID NOs 16 to 110 and to lactose and possibly to (etc. ) any of the other acceptors 2'FL and/or 3FL has α-1,2-fucosyltransferase activity, or (iii) is a functional fragment of any of SEQ ID NOs 16 to 110 and has alpha-1,2-fucosyltransferase activity, or (iv) comprises a polypeptide comprising at least 80% sequence identity to the full-length amino acid sequence of any one of SEQ ID NOs 16 to 110 and b. at least one alpha-1,3-fucosyltransferase, the fucosyltransferase Enzyme (i) comprises as any one of SEQ ID NOs: 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, any one of 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NO: any one of 113, 114, 118, 119, 120, 121 or 122, even better NO: 113 or 114, preferably the polypeptide sequence of SEQ ID NO: 114, or (ii) any one of SEQ ID NO: 111 to 132, preferably any one of SEQ ID NO: 113 to 132, more Preferably any one of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably SEQ ID NOs: 113, 114, 118, 119, 120 , any one of 121 or 122, even more preferably SEQ ID NO: 113 or 114, a functional homologue, variant or derivative of SEQ ID NO: 114, compared with those having SEQ ID NOs 111 to 132, Preferably any one of SEQ ID NOs: 113 to 132, more preferably any one of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even More preferably SEQ ID NOs: 113, 114, 1 Any one of 18, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: 114 in the alpha-1,3-fucosyltransferase polypeptide The full length of either has at least 80% overall sequence identity and has alpha-1,3-fucosylation to lactose and possibly to any of the other receptor(s) 2'FL and/or 3FL Enzymatic activity, or (iii) any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120 , any one of 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NO: any one of 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113,114,118,119,120,121 or 122 ID NO: 113 or 114, the optimal functional fragment of SEQ ID NO: 114 and having alpha-1,3-fucosyltransferase activity, or (iv) comprising a polypeptide comprising the any one, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132 any one, even better any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: 114 The full-length amino acid sequence has or consists of an amino acid sequence with at least 80% sequence identity and has α-1,3-fucosyltransferase activity. 8. The method of specific example 7, wherein the cell expresses: a. at least one alpha-1,2-fucosyltransferase, the fucosyltransferase (i) comprising as SEQ ID NOs: 19 to 21 , any of 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 The polypeptide sequence of either, or (ii) is SEQ ID NO 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 A functional homologue, variant or derivative of any of to 91, 96 to 100, 102, 103 or 105, with SEQ ID NOs 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 of the α-1,2-fucosyltransferase polypeptide The full length of either has at least 80% overall sequence identity and has alpha-1,2-fucosylation to lactose and possibly any of the other receptor(s) 2'FL and/or 3FL Enzymatic activity, or (iii) is SEQ ID NOs 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91 , a functional fragment of any one of 96 to 100, 102, 103, or 105 and having alpha-1,2-fucosyltransferase activity, or (iv) comprising a polypeptide comprising and SEQ ID NO 19 to Any of 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 The full-length amino acid sequence of one has or consists of an amino acid sequence having at least 80% sequence identity and has alpha-1,2-fucosyltransferase activity, and b. at least one alpha-1,3 - a fucosyltransferase, the fucosyltransferase (i) comprising as any one of SEQ ID NOs: 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably Any of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NOs: 113, 114, 118, 119, 120, Any one of 121 or 122, even more preferably the polypeptide sequence of SEQ ID NO: 113 or 114, the best SEQ ID NO: 114, or (ii) is SEQ ID NO 111 Any one of to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or any one of 132, even better any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO : a functional homologue, variant or derivative of 114, with any one of SEQ ID NOs: 111 to 132, preferably SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, any one of 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably any one of SEQ ID NOs: 113, 114, 118, 119, 120, 121 or 122, even More preferably, the full length of any of the alpha-1,3-fucosyltransferase polypeptides of SEQ ID NO: 113 or 114, most preferably SEQ ID NO: 114 has at least 80% overall sequence identity and is consistent with lactose and may have α-1,3-fucosyltransferase activity for any of the (these) other receptors 2'FL and/or 3FL, or (iii) as in SEQ ID NOs 111 to 132 Any one, preferably any one of SEQ ID NOs: 113 to 132, more preferably any one of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132 Any one, even better any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: 114 function fragment and having alpha-1,3-fucosyltransferase activity, or (iv) comprising a polypeptide comprising the any one, more preferably any one of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably SEQ ID NOs: 113, 114 , any one of 118, 119, 120, 121 or 122, even more preferably SEQ ID NO: 113 or 114, the full-length amino acid sequence of SEQ ID NO: 114 has an amino group with at least 80% sequence identity Acid sequence or consisting of and having alpha-1,3-fucosyltransferase activity. 9. The method of specific example 7, wherein the cell expresses: a. at least one alpha-1,2-fucosyltransferase, the fucosyltransferase (i) comprising as SEQ ID NOs: 16 to 19 any one, preferably SEQ ID NO 16 or 19, more preferably the polypeptide sequence of SEQ ID NO 16, or (ii) any one of SEQ ID NO 16 to 19, preferably SEQ ID NO 16 or 19 , preferably a functional homologue, variant or derivative of SEQ ID NO 16, and the alpha-1,2 having SEQ ID NO 16 to 19, preferably SEQ ID NO 16 or 19, more preferably SEQ ID NO 16 - the full length of any of the fucosyltransferase polypeptides has at least 80% overall sequence identity and alpha to lactose and possibly any of the other receptor(s) 2'FL and/or 3FL -1,2-fucosyltransferase activity, or (iii) is any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 or 19, more preferably a functional fragment of SEQ ID NO 16 and has α-1,2-fucosyltransferase activity, or (iv) comprising a polypeptide comprising any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 or 19, more preferably SEQ ID The full-length amino acid sequence of NO 16 has or consists of an amino acid sequence having at least 80% sequence identity and has alpha-1,2-fucosyltransferase activity, and b. at least one alpha-1,3 - a fucosyltransferase, the fucosyltransferase (i) comprising as any one of SEQ ID NOs: 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably Any of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NOs: 113, 114, 118, 119, 120, Any one of 121 or 122, even more preferably the polypeptide sequence of SEQ ID NO: 113 or 114, preferably SEQ ID NO: 114, or (ii) any one of SEQ ID NOs 111 to 132, preferably Any one of SEQ ID NOs: 113 to 132, more preferably any one of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more Preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best functional homologue, variant of SEQ ID NO: 114 or Derivatives with SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129 , any one of 130, 131 or 132, even better SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, most Preferably the full length of any of the alpha-1,3-fucosyltransferase polypeptides of SEQ ID NO: 114 has at least 80% overall sequence identity and is compatible with lactose and possibly other receptors 2 Either 'FL and/or 3FL has alpha-1,3-fucosyltransferase activity, or (iii) is any one of SEQ ID NOs 111 to 132, preferably SEQ ID NO: 113 any of to 132, more preferably any of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NOs: Any one of 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, a functional fragment of best SEQ ID NO: 114 with alpha-1,3-fucoid Glycosyltransferase activity, or (iv) comprising a polypeptide comprising any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: any one of 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably in SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122 Any one, even more preferably SEQ ID NO: 113 or 114, the full-length amino acid sequence of SEQ ID NO: 114 having or consisting of an amino acid sequence having at least 80% sequence identity and having alpha-1 , 3-fucosyltransferase activity. 10. The method of any one of specific examples 4 to 9, wherein the cell produces the lactose for synthesizing any one of the 2'FL, 3-FL and DiFL, preferably the cell further produces the one or more at least one of the other receptors. 11. The method of any one of embodiments 1 to 10, wherein the method comprises using one or more precursors for synthesizing any of the 2'FL, 3-FL and DiFL. 12. The method of any one of embodiments 1 to 11, wherein the precursor(s) and/or acceptors are fully converted into any of the 2'FL, 3-FL and DiFL. 13. The method of any one of embodiments 4 to 12, wherein the cell is modified in the expression or activity of at least one of the fucosyltransferases. 14. The method of any one of embodiments 4 to 13, wherein the cell is a metabolically engineered cell. 15. The method of embodiment 14, wherein the cell comprises multiple copies of the same coding DNA sequence encoding a protein. 16. The method of any one of embodiments 4 to 15, wherein the cell is modified to produce GDP-fucose. 17. The method of any one of embodiments 4 to 16, wherein the cell is modified for enhanced GDP-fucose production compared to an unmodified precursor cell, and wherein the modification is selected from the group consisting of: Group of those: UDP-glucose: deletion of the gene encoding undecyl isopentenyl-phosphoglucose-1-phosphotransferase, overexpression of the gene encoding GDP-L-fucose synthase, GDP-mannose4,6 - Overexpression of a gene encoding dehydratase, overexpression of a gene encoding mannose-1-phosphate guanosyltransferase, overexpression of a gene encoding phosphomannose mutase, or overexpression of a gene encoding mannose-6-phosphate isomerase of excessive performance. 18. The method of any one of embodiments 4 to 17, wherein the cells are resistant to lactose killing when grown in an environment where lactose is combined with one or more other carbon sources. 19. The method of any one of specific examples 4 to 18, wherein the cell intracellularly produces the DiFL or the mixture of the 2'FL, 3-FL and DiFL and wherein the produced DiFL, 2'FL and/or Some or substantially all of the 3-FL remains intracellular and/or is excreted outside the cell via passive or active transport. 20. The method of any one of embodiments 4 to 19, wherein the cell is further genetically modified for i) modified expression of endogenous membrane proteins, and/or ii) modified activity of endogenous membrane proteins, and/or iii) expression of homologous membrane proteins, and/or iv) expression of heterologous membrane proteins, wherein the membrane proteins are involved in the secretion of the 2'FL, 3-FL and DiFL from the mixture to the outside of the cell Either, preferably, the membrane protein is involved in the secretion of all of the 2'FL, 3-FL and DiFL from the mixture to the outside of the cell. 21. The method of any one of embodiments 4 to 20, wherein the cell is further genetically modified for i) modified expression of endogenous membrane proteins, and/or ii) modified activity of endogenous membrane proteins, and/or iii) expression of a homologous membrane protein, and/or iv) expression of a heterologous membrane protein, wherein the membrane protein is involved in the uptake of lactose for the synthesis of any of the 2'FL, 3-FL and DiFL and/or at least one other receptor 2'FL and/or 3FL. 22. The method of any one of embodiment 20 or 21, wherein the membrane protein is selected from a list comprising the following: transporter, PP-bond hydrolysis-driven transporter, β-barrel Porosin, auxiliary transport protein (transport protein), putative transport protein and phosphate transfer driven group translocation protein, preferably, these transport proteins comprise MFS transport protein, sugar efflux transport protein and chelatin export protein , Preferably, the transport proteins driven by hydrolysis of PP-bonds comprise ABC transport proteins and chelatin export proteins. 23. The method of any one of specific examples 20 to 22, wherein the membrane protein improves the production of any one of the 2'FL, 3-FL and DiFL and/or can achieve and/or enhance the 2'FL , outflow of any of 3-FL and DiFL. 24. The method of any one of embodiments 4 to 23, wherein the cells are stably cultured in culture medium. 25. as the method of any one in specific example 4 to 24, wherein these conditions comprise: (i) use comprises at least one for producing this DiFL or this 2'FL, 3-FL and DiFL mixture precursor and/ or acceptor's medium, and/or (ii) adding to the medium at least one precursor and/or acceptor feed for the production of the DiFL or the mixture of 2'FL, 3-FL and DiFL. 26. The method of any one of specific examples 4 to 25, comprising at least one of the following steps: i) using a culture medium comprising at least one precursor and/or acceptor; ii) adding the The medium is supplemented with at least one precursor and/or acceptor feed, wherein the total reactor volume is in the range of 250 mL (milliliters) to 10.000 ^m3 (cubic meters), preferably in a continuous manner, and preferably such that the medium is the final volume does not exceed three times, preferably not more than twice, more preferably less than twice the volume of the medium before adding the precursor and/or acceptor feed; iii) adding at least one precursor to the medium in the reactor Body and/or acceptor feed, wherein the total reactor volume is in the range of 250 mL (milliliters) to 10.000 ^m3 (cubic meters), preferably in a continuous manner, and preferably such that the final volume of the medium is not More than three times, preferably not more than twice, more preferably less than twice the volume of the medium before adding the precursor and/or acceptor feed, and wherein preferably, the precursor and/or acceptor feed is The pH is set between 3 and 7, and wherein preferably the temperature of the precursor and/or acceptor feed is maintained between 20°C and 80°C; iv) over 1 day, 2 days, 3 days, 4 Add at least one precursor and/or acceptor feed to the medium in a continuous manner by means of a feed solution over a time course of 1 day, 5 days; v) over a period of 1 day, 2 days, 3 days, 4 days, 5 days The process adds at least one precursor and/or acceptor feed to the medium in a continuous manner by means of a feed solution, and wherein preferably the pH of the feed solution is set between 3 and 7, and wherein preferably , the temperature of the feed solution is maintained between 20°C and 80°C; the method produces a concentration in the final culture of at least 50 g/L, preferably at least 75 g/L, more preferably at least 90 g/L, more preferably DiFL or the 2'FL, 3-FL and DiFL of at least 100 g/L, more preferably at least 125 g/L, more preferably at least 150 g/L, more preferably at least 175 g/L, more preferably at least 200 g/L Any of the 2'FL, 3-FL and DiFL in the mixture. 27. The method of any one of Examples 4 to 25, comprising at least one of the following steps: i) using a method comprising at least 50, more preferably at least 75, more preferably at least 100, more preferably at least 120, more preferably at least 150 grams of lactose medium, wherein the reactor volume is in the range of 250 mL to 10.000 ^m3 (cubic meters); ii) add to the medium a medium containing at least 50,000,000,000 per liter of initial reactor volume, More preferably at least 75, more preferably at least 100, more preferably at least 120, more preferably at least 150 grams of lactose feed of lactose, wherein the reactor volume is in the range of 250 mL to 10.000 ^m3 (cubic meters), preferably with adding in a continuous manner, and preferably such that the final volume of the medium does not exceed three times, preferably no more than twice, more preferably less than 2 times the volume of the medium before the addition of the lactose feed; iii) adding to the medium contains per liter Lactose feed of initial reactor volume of at least 50, more preferably at least 75, more preferably at least 100, more preferably at least 120, more preferably at least 150 grams of lactose, wherein the reactor volume is between 250 mL and 10.000 m ³ (cubic meters) ), preferably in a continuous manner, and preferably such that the final volume of the medium is no more than three times, preferably no more than two times, more preferably less than two times the volume of the medium before the addition of the lactose feed, and wherein Preferably, the pH of the lactose feed is set between 3 and 7, and wherein preferably the temperature of the lactose feed is maintained between 20°C and 80°C; iv) over 1 day, 2 days, 3 Lactose feed was added to the medium in a continuous manner with the aid of the feed solution over the course of days, 4 days, 5 days; v) over the course of 1 day, 2 days, 3 days, 4 days, 5 days with the aid of the feed The solution adds lactose feed to the medium in a continuous manner, and wherein the lactose feed solution has a concentration of 50 g/L, preferably 75 g/L, more preferably 100 g/L, more preferably 125 g/L, more preferably 150 g/L, better 175 g/L, better 200 g/L, better 225 g/L, better 250 g/L, better 275 g/L, better 300 g/L, better 325 g/L, better 350 g/L, better 375 g/L, better 400 g/L, better 450 g/L, better 500 g/L, even better 550 g/L, best 600 g/L; and wherein preferably, the pH of the feed solution is set between 3 and 7, and wherein preferably, the temperature of the feed solution is maintained between 20°C and 80°C; The resulting concentration in the final volume of the substance is at least 50 g/L, preferably at least 75 g/L, more preferably at least 90 g/L, more preferably at least 100 g/L, more preferably at least 125 g/L, more preferably at least 150 g/L g/L, preferably at least 175 g/L, more preferably at least 200 g/L of DiFL or any of the 2'FL, 3-FL and DiFL in the mixture of 2'FL, 3-FL and DiFL. 28. The method of embodiment 27, wherein the lactose feed is started by culturing at a concentration of at least 5 mM, preferably at a concentration of 30, 40, 50, 60, 70, 80, 90, 100, 150 mM , preferably by adding lactose at a concentration of >300 mM. 29. The method of any one of embodiments 27 or 28, wherein the lactose feed is by adding lactose to the culture at a concentration such that at least 5 mM, preferably 10 is obtained throughout the production phase of the culture. mM or 30 mM lactose concentration. 30. The method of any one of embodiments 4 to 29, wherein the cells are cultured for at least about 60, 80, 100 or about 120 hours or in a continuous manner. 31. The method of any one of specific examples 4 to 30, wherein the cell is cultured in a medium comprising a carbon source, the carbon source comprising monosaccharides, disaccharides, oligosaccharides, polysaccharides, polyols, glycerol; In a complex medium of molasses, corn infusion, egg white, pancreas or yeast extract; preferably, wherein the carbon source is selected from the list comprising the following: glucose, glycerol, fructose, sucrose, maltose, lactose, arabinose , malto-oligosaccharide, maltotriose, sorbitol, xylose, rhamnose, galactose, mannose, methanol, ethanol, trehalose, starch, cellulose, hemicellulose, molasses, corn infusion, High fructose syrup, acetate, citrate, lactate and pyruvate. 32. The method of any one of embodiments 1 to 3, 5, 11 or 12, wherein the cell-free system comprises: a. at least one alpha-1,2-fucosyltransferase, the fucosyl The transferase (i) comprises the polypeptide sequence of any one of SEQ ID NOs: 16 to 110, or (ii) is a functional homologue, variant or derivative of any one of SEQ ID NOs 16 to 110, The functional homologue, variant or derivative has at least 80% overall sequence identity to the full length of any of the alpha-1,2-fucosyltransferase polypeptides having SEQ ID NOs 16-110 and has alpha-1,2-fucosyltransferase activity on lactose and possibly on any of the (these) other receptors 2'FL and/or 3FL, or (iii) as SEQ ID NOs 16 to 110 A functional fragment of any one and having alpha-1,2-fucosyltransferase activity, or (iv) comprising a polypeptide comprising a full-length amine group with any one of SEQ ID NOs 16 to 110 An amino acid sequence having or consisting of an amino acid sequence having at least 80% sequence identity and having alpha-1,2-fucosyltransferase activity, and b. at least one alpha-1,3-fucosyltransferase Enzyme, the fucosyltransferase (i) comprises as any one of SEQ ID NO: 111 to 132, preferably any one of SEQ ID NO: 113 to 132, more preferably SEQ ID NO: 113, any of 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better any of SEQ ID NOs: 113, 114, 118, 119, 120, 121 or 122 One, even more preferably the polypeptide sequence of SEQ ID NO: 113 or 114, preferably SEQ ID NO: 114, or (ii) any one of SEQ ID NOs 111 to 132, preferably SEQ ID NO: 113 to any one of 132, more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NO: 113 , any one of 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, a functional homologue, variant or derivative of SEQ ID NO: 114, with or any one of 132, even better SEQ ID NO : any one of 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, the alpha-1,3-fucosyl of SEQ ID NO: 114 The full-length of any of the transferase polypeptides has at least 80% overall sequence identity and α-1,3- for lactose and possibly for any of the other receptor(s) 2'FL and/or 3FL Fucosyltransferase activity, or (iii) any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, any of 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better any of SEQ ID NOs: 113, 114, 118, 119, 120, 121 or 122 , even more preferably SEQ ID NO: 113 or 114, optimally a functional fragment of SEQ ID NO: 114 and having alpha-1,3-fucosyltransferase activity, or (iv) comprising a polypeptide comprising the Any one of ID Nos 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, Any one of 130, 131 or 132, even better SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best The full-length amino acid sequence of SEQ ID NO: 114 has or consists of an amino acid sequence having at least 80% sequence identity and has alpha-1,3-fucosyltransferase activity, and c. GDP-fucos sugar, and d. at least one of lactose and possibly such other receptor(s). 33. The method of specific example 32, wherein the cell-free system comprises: a. at least one alpha-1,2-fucosyltransferase, the fucosyltransferase (i) comprising as SEQ ID NO: 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 The polypeptide sequence of any one, or (ii) is SEQ ID NOs 19-21, 23-38, 40-44, 46-48, 50-52, 54-61, 64, 65, 69-73, 76-86 A functional homologue, variant or derivative of any of , 88 to 91, 96 to 100, 102, 103 or 105 with SEQ ID NOs 19 to 21, 23 to 38, 40 to 44, 46 to The alpha-1,2-fucosyltransferase polypeptide of 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 The full length of any of them has at least 80% overall sequence identity and has alpha-1,2-fucose for lactose and possibly any of the other receptor(s) 2'FL and/or 3FL Syltransferase activity, or (iii) SEQ ID NOs 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 A functional fragment of any of to 91, 96 to 100, 102, 103 or 105 and having alpha-1,2-fucosyltransferase activity, or (iv) comprising a polypeptide comprising the 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 The full-length amino acid sequence of any one has or consists of an amino acid sequence having at least 80% sequence identity and has alpha-1,2-fucosyltransferase activity, and b. at least one alpha-1 , 3-fucosyltransferase, the fucosyltransferase (i) comprises any one of SEQ ID NOs: 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, More preferably any one of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably SEQ ID NOs: 113, 114, 118, 119, Any one of 120, 121 or 122, even more preferably the polypeptide sequence of SEQ ID NO: 113 or 114, the best SEQ ID NO: 114, or (ii) is SEQ ID NO Any one of 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, Any one of 131 or 132, even better SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID A functional homologue, variant or derivative of NO: 114 with any one of SEQ ID NOs: 111 to 132, preferably SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118 , any one of 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably any one of SEQ ID NOs: 113, 114, 118, 119, 120, 121 or 122, Even more preferably SEQ ID NO: 113 or 114, the full length of any of the alpha-1,3-fucosyltransferase polypeptides of SEQ ID NO: 114 has at least 80% overall sequence identity and is consistent with Lactose and possibly α-1,3-fucosyltransferase activity for any of the (these) other receptors 2'FL and/or 3FL, or (iii) as in SEQ ID NOs 111 to 132 any one, preferably any one of SEQ ID NOs: 113 to 132, more preferably among SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132 any one, even better any of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: 114 A functional fragment and having α-1,3-fucosyltransferase activity, or (iv) comprising a polypeptide comprising any one of SEQ ID NOs: 111 to 132, preferably SEQ ID NOs: 113 to 132 any one, more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably SEQ ID NO: 113, Any one of 114, 118, 119, 120, 121 or 122, even more preferably SEQ ID NO: 113 or 114, the full-length amino acid sequence of SEQ ID NO: 114 has an amine having at least 80% sequence identity base acid sequence or consisting of it and having alpha-1,3-fucosyltransferase activity, and c. GDP-fucose, and at least one of d. lactose and possibly such other receptor(s). 34. The method of specific example 32, wherein the cell-free system comprises: a. at least one alpha-1,2-fucosyltransferase, the fucosyltransferase (i) comprising as SEQ ID NO: 16 any one of to 19, preferably SEQ ID NO 16 or 19, more preferably the polypeptide sequence of SEQ ID NO 16, or (ii) any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 Or 19, more preferably a functional homologue, variant or derivative of SEQ ID NO 16, and the alpha-1 having SEQ ID NO 16 to 19, preferably SEQ ID NO 16 or 19, more preferably SEQ ID NO 16 , the full length of any of the 2-fucosyltransferase polypeptides has at least 80% overall sequence identity and is compatible with lactose and possibly any of the (etc.) other receptors 2'FL and/or 3FL Has α-1,2-fucosyltransferase activity, or (iii) is a functional fragment of any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 or 19, more preferably SEQ ID NO 16 and has α-1,2-fucosyltransferase activity, or (iv) comprises a polypeptide comprising any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 or 19, more preferably The full-length amino acid sequence of SEQ ID NO 16 has or consists of an amino acid sequence having at least 80% sequence identity and has alpha-1,2-fucosyltransferase activity, and b. at least one alpha-1 , 3-fucosyltransferase, the fucosyltransferase (i) comprises any one of SEQ ID NOs: 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, More preferably any one of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably SEQ ID NOs: 113, 114, 118, 119, any one of 120, 121 or 122, even more preferably the polypeptide sequence of SEQ ID NO: 113 or 114, best SEQ ID NO: 114, or (ii) any one of SEQ ID NOs 111 to 132, Preferably any one of SEQ ID NOs: 113 to 132, more preferably any one of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, Even better any of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best functional homologue of SEQ ID NO: 114, Variants or derivatives having SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, any one of 128, 129, 130, 131 or 132, even more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even more preferably SEQ ID NO: 113 or 114. The full length of any one of the alpha-1,3-fucosyltransferase polypeptides of optimal SEQ ID NO: 114 has at least 80% overall sequence identity and is identical to lactose and possibly the (etc.) other Either one of the acceptors 2'FL and/or 3FL has alpha-1,3-fucosyltransferase activity, or (iii) is any one of SEQ ID NOs 111 to 132, preferably SEQ ID NO: any one of 113 to 132, more preferably SEQ ID NO: any one of 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better Any of ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, the best functional fragment of SEQ ID NO: 114 with alpha-1,3 - Fucosyltransferase activity, or (iv) comprising a polypeptide comprising any of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs Any of ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NOs: 113, 114, 118, 119, 120, 121 or any one of 122, even more preferably SEQ ID NO: 113 or 114, the full-length amino acid sequence of SEQ ID NO: 114 has or consists of an amino acid sequence having at least 80% sequence identity and has α-1,3-fucosyltransferase activity, and at least one of c. GDP-fucose, and d. lactose and possibly the other receptor(s). 35. The method of any one of specific examples 1 to 34, wherein the separation comprises at least one of the following steps: clarification, ultrafiltration, nanofiltration, two-phase partitioning, reverse osmosis, microfiltration, activated carbon or carbon treatment , treatment with nonionic surfactants, enzymatic digestion, tangential flow high performance filtration, tangential flow ultrafiltration, affinity chromatography, ion exchange chromatography, hydrophobic interaction chromatography and/or gel filtration, ligands Exchange chromatography. 36. The method of any one of specific examples 1 to 35, further comprising purifying any one of the 2'FL, 3-FL or DiFL, preferably all of the 2'FL, 3-FL or DiFL . 37. The method of embodiment 36, wherein the purification comprises at least one of the following steps: using activated carbon or carbon; using charcoal, nanofiltration, ultrafiltration, electrophoresis, enzymatic treatment or ion exchange; using alcohol; using hydroalcoholic Mixtures; Crystallization; Evaporation; Precipitation; Drying, Spray Drying, Freeze Drying, Spray Freeze Drying, Freeze Spray Drying, Band Drying, Belt Drying, Vacuum Belt Drying, Vacuum Belt Drying Drying, tumble drying, tumble drying, vacuum tumble drying or vacuum tumble drying. 38. The method of any one of embodiments 1 to 37, wherein the DiFL is measured by the total amount of 2'FL, 3-FL and DiFL produced in the 2'FL, 3-FL and DiFL mixture. At least 80% purity was purified from the 2'FL, 3-FL and DiFL mixture. 39. A metabolically engineered cell for the production of DiFL or a mixture of DiFL, 2'FL and/or 3-FL, wherein the cell: a. expresses at least two fucosyltransferases, the rocks A fucosyltransferase is capable of transferring a fucose residue from a GDP-fucose donor to any one or more of the acceptor lactose and acceptor 2'FL and/or 3-FL, and b. capable of Synthesis of GDP-fucose, wherein the GDP-fucose is a donor for the fucosyltransferases, c. preferably wherein the cells are metabolically engineered for the production of DiFL or DiFL, 2 A mixture of 'FL and/or 3-FL. 40. The cell of embodiment 39, wherein the cell is modified with at least one gene expression module, characterized in that the expression from any one of these expression modules is persistent or produced by a natural inducer, compared to Preferably continuous. 41. The cell of embodiment 40, wherein the cell comprises multiple copies of the same coding DNA sequence encoding a protein. 42. The cell of any one of embodiments 30 and 31, wherein the cell expresses: a. at least one α-1,2-fucosyltransferase, the fucosyltransferase (i) comprising as SEQ The polypeptide sequence of any one of ID NOs: 16 to 110, or (ii) is a functional homologue, variant or derivative of any one of SEQ ID NOs 16 to 110, the functional homologue, variant or derivatives having at least 80% overall sequence identity to the full length of any of the alpha-1,2-fucosyltransferase polypeptides having SEQ ID NOs 16 to 110 and to lactose and possibly to (etc. ) any of the other acceptors 2'FL and/or 3FL has α-1,2-fucosyltransferase activity, or (iii) is a functional fragment of any of SEQ ID NOs 16 to 110 and has alpha-1,2-fucosyltransferase activity, or (iv) comprises a polypeptide comprising at least 80% sequence identity to the full-length amino acid sequence of any one of SEQ ID NOs 16 to 110 and b. at least one alpha-1,3-fucosyltransferase, the fucosyltransferase Enzyme (i) comprises as any one of SEQ ID NOs: 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, any one of 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NO: any one of 113, 114, 118, 119, 120, 121 or 122, even better NO: 113 or 114, preferably the polypeptide sequence of SEQ ID NO: 114, or (ii) any one of SEQ ID NO: 111 to 132, preferably any one of SEQ ID NO: 113 to 132, more Preferably any one of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably SEQ ID NOs: 113, 114, 118, 119, 120 , any one of 121 or 122, even more preferably SEQ ID NO: 113 or 114, a functional homologue, variant or derivative of SEQ ID NO: 114, compared with those having SEQ ID NOs 111 to 132, Preferably any one of SEQ ID NOs: 113 to 132, more preferably any one of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even More preferably SEQ ID NO: 113, 114 , any one of 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: 114 in the alpha-1,3-fucosyltransferase polypeptide The full length of any one has at least 80% overall sequence identity and has an alpha-1,3-fucosyl group for lactose and possibly any of the other receptor(s) 2'FL and/or 3FL transferase activity, or (iii) any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, any one of 120, 121, 122, 128, 129, 130, 131 or 132, even better any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, the optimal functional fragment of SEQ ID NO: 114 and having alpha-1,3-fucosyltransferase activity, or (iv) comprising a polypeptide comprising SEQ ID NO: 111 to any one of 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or Any one of 132, even better any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: The full-length amino acid sequence of 114 has or consists of an amino acid sequence of at least 80% sequence identity and has alpha-1,3-fucosyltransferase activity. 43. The cell of specific example 42, wherein the cell expresses: a. at least one alpha-1,2-fucosyltransferase, the fucosyltransferase (i) comprising as SEQ ID NOs: 19 to 21 , any of 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 The polypeptide sequence of either, or (ii) is SEQ ID NO 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 A functional homologue, variant or derivative of any of to 91, 96 to 100, 102, 103 or 105, with SEQ ID NOs 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 of the α-1,2-fucosyltransferase polypeptide The full length of either has at least 80% overall sequence identity and has alpha-1,2-fucosylation to lactose and possibly any of the other receptor(s) 2'FL and/or 3FL Enzymatic activity, or (iii) is SEQ ID NOs 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91 , a functional fragment of any one of 96 to 100, 102, 103, or 105 and having alpha-1,2-fucosyltransferase activity, or (iv) comprising a polypeptide comprising and SEQ ID NO 19 to Any of 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 The full-length amino acid sequence of one has or consists of an amino acid sequence having at least 80% sequence identity and has alpha-1,2-fucosyltransferase activity, and b. at least one alpha-1,3 - a fucosyltransferase, the fucosyltransferase (i) comprising as any one of SEQ ID NOs: 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably Any of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NOs: 113, 114, 118, 119, 120, Any one of 121 or 122, even more preferably the polypeptide sequence of SEQ ID NO: 113 or 114, the best SEQ ID NO: 114, or (ii) is SEQ ID NO 11 Any one of 1 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, Any one of 131 or 132, even better SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID A functional homologue, variant or derivative of NO: 114 with any one of SEQ ID NOs: 111 to 132, preferably SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118 , any one of 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably any one of SEQ ID NOs: 113, 114, 118, 119, 120, 121 or 122, Even more preferably SEQ ID NO: 113 or 114, the full length of any of the alpha-1,3-fucosyltransferase polypeptides of SEQ ID NO: 114 has at least 80% overall sequence identity and is consistent with Lactose and possibly α-1,3-fucosyltransferase activity for any of the (these) other receptors 2'FL and/or 3FL, or (iii) as in SEQ ID NOs 111 to 132 any one, preferably any one of SEQ ID NOs: 113 to 132, more preferably among SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132 any one, even better any of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: 114 A functional fragment and having α-1,3-fucosyltransferase activity, or (iv) comprising a polypeptide comprising any one of SEQ ID NOs: 111 to 132, preferably SEQ ID NOs: 113 to 132 any one, more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably SEQ ID NO: 113, Any one of 114, 118, 119, 120, 121 or 122, even more preferably SEQ ID NO: 113 or 114, the full-length amino acid sequence of SEQ ID NO: 114 has an amine having at least 80% sequence identity The amino acid sequence or consists of it and has α-1,3-fucosyltransferase activity. 44. The cell of specific example 42, wherein the cell expresses: a. at least one alpha-1,2-fucosyltransferase, the fucosyltransferase (i) comprising as SEQ ID NOs: 16 to 19 any one, preferably SEQ ID NO 16 or 19, more preferably the polypeptide sequence of SEQ ID NO 16, or (ii) any one of SEQ ID NO 16 to 19, preferably SEQ ID NO 16 or 19 , preferably a functional homologue, variant or derivative of SEQ ID NO 16, and the alpha-1,2 having SEQ ID NO 16 to 19, preferably SEQ ID NO 16 or 19, more preferably SEQ ID NO 16 - the full length of any of the fucosyltransferase polypeptides has at least 80% overall sequence identity and alpha to lactose and possibly any of the other receptor(s) 2'FL and/or 3FL -1,2-fucosyltransferase activity, or (iii) is any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 or 19, more preferably a functional fragment of SEQ ID NO 16 and has α-1,2-fucosyltransferase activity, or (iv) comprising a polypeptide comprising any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 or 19, more preferably SEQ ID The full-length amino acid sequence of NO 16 has or consists of an amino acid sequence having at least 80% sequence identity and has alpha-1,2-fucosyltransferase activity, and b. at least one alpha-1,3 - a fucosyltransferase, the fucosyltransferase (i) comprising as any one of SEQ ID NOs: 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably Any of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NOs: 113, 114, 118, 119, 120, Any one of 121 or 122, even more preferably the polypeptide sequence of SEQ ID NO: 113 or 114, preferably SEQ ID NO: 114, or (ii) any one of SEQ ID NOs 111 to 132, preferably Any one of SEQ ID NOs: 113 to 132, more preferably any one of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more Preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best functional homologue, variant of SEQ ID NO: 114 or derivatives, with SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, any one of 129, 130, 131 or 132, even more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even more preferably SEQ ID NO: 113 or 114, The full length of any of the alpha-1,3-fucosyltransferase polypeptides of optimal SEQ ID NO: 114 has at least 80% overall sequence identity and is compatible with lactose and possibly other receptor(s) Any one of 2'FL and/or 3FL has alpha-1,3-fucosyltransferase activity, or (iii) is any one of SEQ ID NOs 111 to 132, preferably SEQ ID NO: any one of 113 to 132, more preferably any one of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better : any one of 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, a functional fragment of best SEQ ID NO: 114 with alpha-1,3-rock Fluorosyltransferase activity, or (iv) comprising a polypeptide comprising any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NO : any one of 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122 Any one, even more preferably SEQ ID NO: 113 or 114, the full-length amino acid sequence of SEQ ID NO: 114 having or consisting of an amino acid sequence having at least 80% sequence identity and having an α- 1,3-fucosyltransferase activity. 45. The cell of any one of specific examples 39 to 44, wherein the cell produces the lactose for synthesizing any one of the 2'FL, 3-FL and DiFL, preferably the cell further produces the one or more at least one of the other receptors. 46. The cell of any one of embodiments 39 to 45, wherein the cell uses one or more precursors for synthesizing any one of the 2'FL, 3-FL and DiFL. 47. The cell of any one of embodiments 39 to 46, wherein the precursor(s) and/or receptors are fully converted to any of the 2'FL, 3-FL and DiFL. 48. The cell of any one of embodiments 39 to 47, wherein the cell is modified in the expression or activity of at least one of the fucosyltransferases. 49. The cell of any one of embodiments 39 to 48, wherein the cell is modified to produce GDP-fucose. 50. The cell of any one of embodiments 39 to 49, wherein the cell is modified for enhanced GDP-fucose production compared to an unmodified precursor cell, and wherein the modification is selected from the group consisting of: Group of those: UDP-glucose: deletion of the gene encoding undecyl isopentenyl-phosphoglucose-1-phosphotransferase, overexpression of the gene encoding GDP-L-fucose synthase, GDP-mannose4,6 - Overexpression of a gene encoding dehydratase, overexpression of a gene encoding mannose-1-phosphate guanosyltransferase, overexpression of a gene encoding phosphomannose mutase, or overexpression of a gene encoding mannose-6-phosphate isomerase of excessive performance. 51. The cell of any one of embodiments 39 to 50, wherein the cell is resistant to lactose killing when grown in an environment where lactose is combined with one or more other carbon sources. 52. The cell of any one of specific examples 39 to 51, wherein the cell intracellularly produces the DiFL or the mixture of the 2'FL, 3-FL and DiFL and wherein the produced DiFL, 2'FL and/or Some or substantially all of the 3-FL remains intracellular and/or is excreted outside the cell via passive or active transport. 53. The cell of any one of specific examples 39 to 52, wherein the cell is further genetically modified for i) modified expression of endogenous membrane proteins, and/or ii) modified activity of endogenous membrane proteins, and/or iii) expression of homologous membrane proteins, and/or iv) expression of heterologous membrane proteins, wherein the membrane proteins are involved in the secretion of the 2'FL, 3-FL and DiFL from the mixture to the outside of the cell Either, preferably, the membrane protein is involved in the secretion of all of the 2'FL, 3-FL and DiFL from the mixture to the outside of the cell. 54. The cell of any one of specific examples 39 to 53, wherein the cell is further genetically modified for i) modified expression of endogenous membrane proteins, and/or ii) modified activity of endogenous membrane proteins, and/or iii) expression of a homologous membrane protein, and/or iv) expression of a heterologous membrane protein, wherein the membrane protein is involved in the uptake of lactose for the synthesis of any of the 2'FL, 3-FL and DiFL and/or at least one other receptor 2'FL and/or 3FL. 55. The cell of any one of specific examples 53 or 54, wherein the membrane protein is selected from a list comprising: transporter, PP-bond hydrolysis-driven transporter, β-barrel porin, assisted transport Proteins, putative transport proteins and phosphate transfer driven group translocation proteins, preferably, these transport proteins comprise MFS transport proteins, sugar efflux transport proteins and chelating ferritin export proteins, preferably, these PP- Bond hydrolysis-driven transport proteins include ABC transport proteins and chelatin export proteins. 56. The cell of any one of specific examples 53 to 55, wherein the membrane protein improves the production of any one of the 2'FL, 3-FL and DiFL and/or can achieve and/or enhance the 2'FL , outflow of any of 3-FL and DiFL. 57. The cell of any one of specific examples 39 to 56 or the method of any one of specific examples 4 to 31 and 35 to 38, wherein the cell is a bacterium, a fungus, a yeast, a plant cell, an animal cell or a progenitor. worm cells, - preferably, the bacteria are Escherichia coli ( Escherichia coli ) strains, more preferably Escherichia coli strains of K-12 strains, even more preferably the Escherichia coli K-12 strains are Escherichia coli MG1655, - preferably , the fungus belongs to a genus selected from the group comprising Rhizopus , Dictyostelium , Penicillium , Mucor or Aspergillus , - preferably, the yeast belongs to a genus selected from the group consisting of: Saccharomyces , Zygosaccharomyces , Pichia , Komagataella , Han Hansenula , Yarrowia , Starmerella , Kluyveromyces or Debaromyces , - preferably, the plant cell are algal cells or derived from tobacco, alfalfa, rice, tomato, cotton, rapeseed, soybean, maize or corn plants, - preferably, the animal cells are derived from non-human mammals, birds, fish, invertebrates, reptiles Organoids, amphibians or insects, or genetically modified cell lines derived from human cells excluding embryonic stem cells, more preferably the human and non-human mammalian cells are epithelial cells, embryonic kidney cells, fibroblasts, COS cells , Chinese hamster ovary (Chinese hamster ovary; CHO) cells, murine myeloma cells, NIH-3T3 cells, non-mammary adult stem cells or derivatives thereof, preferably the insect cells are derived from Spodoptera frugiperda , Bombyx mori , Mamestra brassicae , Trichoplusia ni or Drosophila melanogaster , - preferably, the protozoal cell is Leishmania dow ( Leishmania tarentolae ) cells. 58. The cell of specific example 57 or the method of specific example 57, wherein the cell is a surviving Gram-negative bacterium comprising a reduction or elimination compared to unmodified precursor cells Synthetic poly-N-acetyl-glucosamine (poly-N-acetyl-glucosamine; PNAG), Enterobacterial Common Antigen (ECA), cellulose, kolac acid, core oligosaccharide, osmotic regulation Osmoregulated Periplasmic Glucan (OPG), glycerol glucoside, polysaccharide and/or trehalose. 59. A cell such as any one of specific examples 39 to 58 or use of the method of any one of specific examples 1 to 38, 57 and 58 for the production of DiFL, 2'FL and/or 3-FL of the mixture. 60. A method for producing 2',3-difucosyllactose (DiFL), the method comprising the steps of: i) providing the cell of any one of specific examples 39 to 58, and, ii) in culturing the cells under conditions permitting expression of the fucosyltransferases and synthesis of the GDP-fucose, iii) preferably, isolating at least one of the 2'FL, 3-FL or DiFL from the culture If so, it is better to isolate DiFL from the culture, and even better to isolate all of the 2'FL, 3-FL and DiFL from the culture. 61. Use of the cell of any one of specific examples 39 to 58 for the production of DiFL.

The invention will be described in more detail in the examples.

The following examples serve to further illustrate and illustrate the invention and are not intended to be limiting.

Example Example 1. Materials and Methods for Escherichia coli culture medium

Luria Broth (LB) medium was composed of 1% pancreas (Difco, Erembodegem, Belgium), 0.5% yeast extract (Difco) and 0.5% sodium chloride ( VWR, Leuven, Belgium). Minimal medium in 96-well dishes or shake flasks used in culture experiments contained 2.00 g/L NH4Cl, 5.00 g/L (NH4)2SO4, 2.993 g/L KH2PO4, 7.315 g/L K2HPO4, 8.372 g/L MOPS, 0.5 g/L NaCl, 0.5 g/L MgSO4.7H2O, 30 g/L sucrose or 30 g/L glycerol, 1 ml/L vitamin solution, 100 µl/L molybdate solution, and 1 mL/L selenium solution. As specified in the individual examples, 20 g/L lactose was additionally added to the medium as a precursor. Minimal medium was set to pH 7 using 1M KOH. The vitamin solution is composed of 3.6 g/L FeCl2.4H2O, 5 g/L CaCl2.2H2O, 1.3 g/L MnCl2.2H2O, 0.38 g/L CuCl2.2H2O, 0.5 g/L CoCl2.6H2O, 0.94 g/L ZnCl2, 0.0311 g/L H3BO4, 0.4 g/L Na2EDTA.2H2O and 1.01 g/L thiamine.HCl. The molybdate solution contained 0.967 g/L NaMoO4.2H2O. The selenium solution contained 42 g/L Seo2.

The basic medium for fermentation contains 6.75 g/L NH4Cl, 1.25 g/L (NH4)2SO4, 2.93 g/L KH2PO4 and 7.31 g/L KH2PO4, 0.5 g/L NaCl, 0.5 g/L MgSO4.7H2O, 30 g/L sucrose or 30 g/L glycerol, 1 mL/L vitamin solution, 100 µL/L molybdate solution, and 1 mL/L selenium solution with the same composition as described above. 20 g/L lactose, 2'FL and/or 3-FL were additionally added to the medium as acceptors as specified in the respective examples.

The complex medium was sterilized by autoclaving (121°C, 21 min) and the minimal medium by filtration (0.22 µm Sartorius). If necessary, make the medium selective by adding antibiotics: e.g. chloramphenicol (20 mg/L), carbenicillin (100 mg/L), spectinomycin (40 mg/L) and/or Mycin (50 mg/L). plastid

pKD46 (red helper plastid, ampicillin resistance), pKD3 (contains FRT flanked by chloramphenicol resistance (cat) gene), pKD4 (contains FRT flanked by kanamycin resistance (kan) gene) and The pCP20 (expressing FLP recombinase activity) plasmid system was obtained from Prof. R. Cunin (Vrije Universiteit Brussel, Belgium, in 2007). Plastids were maintained in host E. coli DH5α (F ⁻ , phi80d lacZΔM15 , Δ( lacZYA − argF ) U169, deoR , recA1 , endA1 , hsdR17(rk ⁻ , mk ⁺ ), phoA , supE44 , λ ⁻ , purchased from Invitrogen , thi -1, gyrA96 , rel A1). Strains and Mutations

Escherichia coli K12 MG1655 [λ ^- , F ^- , rph-1] was obtained from Escherichia coli Gene Reserve Center (USA) in March 2007, CGSC strain number: 7740. Gene disruption, gene introduction and gene replacement were performed using techniques disclosed by Datsenko and Wanner (PNAS 97 (2000), 6640-6645). This technique is based on antibiotic selection following homologous recombination by λ Red recombinase. Subsequent catalysis by flippase recombinase ensures removal of the antibiotic selection cassette in the final production strain. Transformants carrying the red helper plastid pKD46 were grown at 30°C in 10 mL of LB medium with ampicillin (100 mg/L) and L-arabinose (10 mM) to an OD ₆₀₀ nm of 0.6. Cells were made electrocompetent by washing with 50 mL of ice-cold water a first time and 1 mL of ice-cold water a second time. Next, cells were resuspended in 50 µL of ice-cold water. 50 µL of cells and 10-100 ng of linear double-stranded DNA product were electroporated by using a Gene Pulser™ (BioRad) (600 Ω, 25 µFD and 250 volts). After electroporation, cells were added to 1 mL of LB medium incubated at 37°C for 1 h and finally spread onto LB agar containing 25 mg/L chloramphenicol or 50 mg/L kanamycin for selection Antibiotic-resistant transformants. Selected mutants were verified by PCR with primers upstream and downstream of the modified region, and grown in LB agar at 42°C for loss of helper plastids. Mutants were tested for ampicillin sensitivity. Linear ds-DNA amplicons were obtained by PCR using pKD3, pKD4 and their derivatives as templates. The primers used have a part of the sequence complementary to the template and another part complementary to the side of the chromosomal DNA on which recombination must occur. For gene body knockouts, homology regions were designed 50-nt upstream and 50-nt downstream of the start and stop codons of the gene of interest. For gene body embedding, the transcription start point (+1) must be considered. PCR products were PCR purified, digested with Dpnl, repurified from agarose gels, and suspended in elution buffer (5 mM Tris, pH 8.0). Selected mutants were transformed with pCP20 plastids, which are heat-induced ampicillin and chloramphenicol-resistant plastids showing temperature-sensitive replication and FLP synthesis. Ampicillin resistant transformants were selected at 30°C, followed by purification of a few colonies in LB at 42°C, and then tested for all antibiotic resistance and loss of FLP helper plastids. Knockout and insertion were checked with control primers.

In one example of GDP-fucose production, the mutant strain is derived from E. coli K12 MG1655, which comprises deletion of the E. coli wcaJ , thyA , lacZ , lacY and lacA genes and gene body insertion of the persistent transcription unit These persistent transcription units contain a sucrose transporter such as, for example, CscB from Escherichia coli W with SEQ ID NO 01, such as, for example, Frk derived from Zymomonas mobilis with SEQ ID NO 02 Fructokinase, such as eg sucrose phosphorylase derived from BaSP of Bifidobacterium adolescentis with SEQ ID NO 03 and lactose permease such as eg lacY derived from Escherichia coli with SEQ ID NO 12. For the production of fucosylated lactose structures, mutant GDP-fucose producing strains are additionally modified with expression plastids comprising a persistent transcription unit for a selectable marker such as, for example, having SEQ ID NO 04 E. coli thyA, as listed in Tables 1 and 2 selected from one or more alpha-1,2-fucosyltransferases and one or more alpha-1,3-fucosylases of SEQ ID NOs 16 to 132 transferase. The persistent transcription unit of the fucosyltransferase can also be present in mutant E. coli strains via gene body insertion. As described in WO2016075243 and WO2012007481, GDP-fucose production can be further knocked out in mutant E. coli strains by genomic knockout of E. coli genes comprising glgC , agp , pfkA , pfkB , pgi , arcA , iclR , pgi and lon optimize. GDP-fucose production can additionally be optimized, including for mannose-6-phosphate isomerase (eg, E. coli manA with SEQ ID NO 05), phosphomannose mutase (eg, with SEQ ID NO 05) manB of ID NO 06), mannose-1-phosphate guanosyltransferase (as e.g. manC having SEQ ID NO 07), GDP-mannose 4,6-dehydratase (as e.g. having SEQ ID NO 08 gmd) and a gene body insertion of a continuous transcription unit of GDP-L-fucose synthase (eg, fcl with SEQ ID NO 09). GDP-fucose production can also be obtained by gene body deletion of the E. coli fucK and fucI genes and gene body insertion of persistent transcription units containing fucose permeases, such as for example from those with SEQ ID fucP of E. coli NO 10, and bifunctional fucokinase/fucose-1-phosphate guanosyltransferase, such as eg fkp from Bacteroides fragilis with SEQ NO ID 11.

In one embodiment of the production of lactose (Gal-b1,4-Glc), the mutant strain is derived from E. coli K12 MG1655 and genetically knocked out with the E. coli lacZ and glk genes and the E. coli galETKM operon and a persistent transcription unit The gene body insertion modification of these persistent transcription units is used as, for example, N-acetylglucosamine beta-1,4- from IgtB of N. meningitidis with SEQ ID NO 13 Galactosyltransferase and UDP-glucose 4-epimerase such as eg galE from E. coli with SEQ ID NO 14.

Alternatively and/or additionally, the production of GDP-fucose and/or fucosylated lactose structures can be further optimized in mutant E. coli strains by gene body insertion of a persistent transcription unit comprising a membrane protein such as, for example, MdfA (UniProt ID A0A2T7ANQ9) from Cronobacter muytjensii , MdfA (UniProt ID D4BC23) from Citrobacter younga , MdfA (UniProt ID P0AEY8) from Escherichia coli , MdfA (UniProt ID G9Z5F4) from Yokenella regensburgei , iceT (UniProt ID A0A024L207) from Escherichia coli, or iceT (UniProt ID D4B8A6) from Citrobacter japonicus.

Preferably, but not necessarily, glycosyltransferases, proteins involved in nucleotide-activated sugar synthesis, and/or membrane proteins are N-terminally and/or C-terminally fused to soluble enhancer tags such as, for example, SUMO tags, MBP tags, His, FLAG, Strep-II, Halo-tag, NusA, thioredoxin, GST and/or Fh8 tags to enhance their solubility (Costa et al., Front. Microbiol. 2014, https://doi.org/10.3389/ fmicb. 2014.00063; Fox et al, Protein Sci. 2001, 10(3), 622-630; Jia and Jeaon, Open Biol. 2016, 6: 160196).

Optionally, mutant E. coli strains are modified with gene body insertions encoding chaperone proteins, such as, for example, DnaK, DnaJ, GrpE or the persistent transcription unit of the GroEL/ES chaperone system (Baneyx F., Palumbo J.L. (2003) Improving Heterologous Protein Folding via Molecular Chaperone and Foldase Co-Expression. In: Vaillancourt P.E. (ed.) E. coliGene Expression Protocols. Methods in Molecular Biology™, Vol. 205. Humana Press).

Optionally, the mutant E. coli strain is modified to produce a glycosyl-minimized E. coli strain comprising a gene body deletion of any one or more of the non-essential glycosyltransferase genes, the non-essential glycosyltransferase genes Contains pgaC, pgaD, rfe, rffT, rffM, bcsA, bcsB, bcsC, wcaA, wcaC, wcaE, wcaI, wcaJ, wcaL, waaH, waaF, waaC, waaU, waaZ, waaJ, waaO, waaB, waaS, waaG, waaQ , wbbl, arnC, arnT, yfdH, wbbK, opgG, opgH, ycjM, glgA, glgB, malQ, otsA, and yaiP.

All persistent promoter, UTR and terminator sequences were derived from Mutalik et al. (Nat. Methods 2013, No. 10, 354-360) and Cambray et al. (Nucleic Acids Res. 2013, 41(9), 5139-5148) Libraries described: The gene lines were expressed using the following promoters: MutalikP5 ("PROM0005_MutalikP5"), MutalikP6 ("PROM0006_MutalikP6"), MutalikP9 (“PROM0009_MutalikP9”), MutalikP10 (“PROM0010_MutalikP10)”, MutalikP12 (“PROM0012_MutalikP12)”, apFAB142 (“PROM0031_apFAB142”) and apFAB146 (“PROM0032_apFAB146”) and by De Mey et al. 34), 1-14) described promoter P14 ("PROM0016_P14"); as described by Mutalik et al. (Nat. Methods 2013, No. 10, 354-360) UTR Gene10-LeuAB-BCD2 (" UTR0002_Gene10-LeuAB-BCD2」）、OmpC_BCD5（「UTR0007_OmpC_BCD5」）、Gene10_LeuL（「UTR0011_Gene10_LeuL」）、ThrA_BCD2（「UTR0013_ThrA_BCD2」）、OmpC_BCD18（「UTR0017_OmpC_BCD18」）、GalE_IptFG（「UTR0038_GalE_IptFG」）及如藉由De Mey等人(BMC Biotechnol. 2007, 4(34), 1-14) as described in UTR P14_UTR ("UTR0019_P14_UTR"); and as described by Kim and Lee (FEBS Letters 1997, 407(3), 353-356) The terminator rnpB_T1 ("TER0004_rnpB_T1"), T7early ("TER0010_T7early") as described by Dunn et al. (Nucleic Acids Res. 1980, 8(10), 2119-2132) and as described by Chen L3S2P53 ("TER0015_L3S2P53") described by et al. (Nat. Methods 2013, 10(7), 659-664).

All genes were sequenced synthetically on Twist Bioscience (twistbioscience.com) or IDT (eu.idtdna.com) and codon usage was adapted using the supplier's tools. The SEQ ID NOs described in the present invention are summarized in Tables 1, 2 and 3.

All strains were stored in cryovials (overnight LB cultures mixed with 70% glycerol at a 1:1 ratio) at -80°C. Table 1 : Summary of α-1,2-fucosyltransferases of the present invention SEQ ID NO organism source Country of Origin of Digital Serial Information 16 Helicobacter pylori synthesis Australia 17 Helicobacter sp. MIT 01-6242 synthesis Australia 18 Akkermansia muciniphila synthesis U.S. 19 Porphyromonas catoniae synthesis unknown 20 Capnocytophaga canis synthesis unknown twenty one Capnocytophaga leadbetteri synthesis Finland twenty two Helicobacter sp. MIT 01-6242 synthesis U.S. twenty three Kingella denitrificans synthesis unknown twenty four Pseudoalteromonas distincta synthesis unknown 25 Pyrromonas catarrhalis F0037 synthesis unknown 26 Campylobacter mucosalis synthesis U.K. 27 Pedobacter kyungheensis synthesis South Korea 28 Autotrophic Noviherbaspirillum autotrophicum synthesis Japan 29 Microbacterium trichothecenolyticum synthesis unknown 30 Candidatus Symbiothrix dinenymphae synthesis Japan 31 Geobacter species synthesis Switzerland 32 Chryseobacterium species synthesis Switzerland 33 Microbacterium species synthesis Germany 34 Water Bear (Ramazzottius varieornatus) synthesis Japan 35 Candidatus Wolfebacteria bacterium synthesis U.S. 36 Candidatus Yanofskybacteria bacterium synthesis U.S. 37 Spirochaetes bacterium synthesis U.S. 38 Pedobacter soli synthesis South Korea 39 Bacteroidales KHT7 synthesis unknown 40 Butyrivibrio sp. TB synthesis unknown 41 Butyrivibrio fibrisolvens synthesis unknown 42 Candidatus Magasanikbacteria bacterium synthesis U.S. 43 Vibrio cellolyticus butyricum synthesis unknown 44 Chryseobacterium scophthalmum synthesis U.K. 45 Akkermansia sp. 54_46 synthesis unknown 46 Spade-shaped sea bean sprouts (Lingula unguis) synthesis Japan 47 spatula sea bean sprouts synthesis Japan 48 Species of the genus "Flexibacter" synthesis U.S. 49 Bacteria of Bacteroidetes synthesis U.S. 50 Geobacter sp. AJM synthesis U.S. 51 Candidatus Planktophila lacus synthesis Switzerland 52 Rhizobacteria PAR1 synthesis U.K. 53 Helicobacter sp. 11S02629-2 synthesis Netherlands 54 Candidate Stud synthesis U.S. 55 Candidate Stud synthesis U.S. 56 Diaminobutyricimonas aerilata synthesis South Korea 57 Flavobacterium magnum synthesis South Korea 58 Litoreibacter ponti synthesis South Korea 59 Dysgonomonas alginatilytica synthesis Japan 60 Campylobacter hyointestinalis subsp. lawsonii synthesis U.S. 61 Phycisphaerales bacterium synthesis unknown 62 Akkermansia sp. synthesis unknown 63 Helicobacter sp. MIT 17-337 synthesis U.S. 64 Geobacter sp. G11 synthesis U.S. 65 Empedobacter brevis synthesis unknown 66 Akkermansia muciniphila synthesis U.S. 67 Akkermansia muciniphila synthesis U.S. 68 Akkermansia muciniphila synthesis U.S. 69 Chitinophagaceae bacterium synthesis U.K. 70 Bacteria of the genus Chitinophaga synthesis U.K. 71 Proteobacteria synthesis U.K. 72 Flaviaesturariibacter sp. 17J68-12 synthesis South Korea 73 Geobacter sp. AR-2-6 synthesis Norway, Spitzbergen 74 Helicobacter jaachi synthesis U.S. 75 Helicobacter japonicus synthesis U.S. 76 Klebsiella pneumoniae synthesis unknown 77 Macrophages phagocytose Dysgonomonas capnocytophagoides synthesis U.S. 78 Flavobacterium species synthesis U.K. 79 Geobacter species synthesis U.K. 80 Hoeflea phototrophica synthesis Germany 81 Subdoligranulum variabile synthesis Denmark 82 Pacific Oyster (Crassostrea gigas) synthesis unknown 83 Pacific Oyster synthesis unknown 84 uncultured bacteria synthesis U.S. 85 Clostridium sp. CAG:510 synthesis unknown 86 Robusta leeches (Helobdella robusta) synthesis U.S. 87 Escherichia coli synthesis U.S. 88 Francisella sp. FSC1006 synthesis Italy 89 Spirosoma radiotolerans synthesis South Korea 90 Radiation resistant helix synthesis South Korea 91 Flavobacterium johnsoniae synthesis U.S. 92 Helicobacter pylori synthesis Australia 93 Helicobacter pylori synthesis Australia 94 Helicobacter mustelae synthesis U.S. 95 Helicobacter pylori synthesis Australia 96 Prevotella melaninogenica synthesis unknown 97 Clostridium bolteae synthesis unknown 98 Clostridiales (Clostridiales) bacteria VE202-29 synthesis Japan 99 Candidate species of swamp methane bacteria (Candidatus Methanosphaerula palustris) E1-9c synthesis U.S. 100 Tannerella sp. CAG: 118 synthesis unknown 101 Bacteroides caccae synthesis unknown 102 Vibrio butyricum species synthesis unknown 103 Parabacteroides johnsonii synthesis Japan 104 Akkermansia muciniphila synthesis U.S. 105 Salmonella enterica synthesis U.S. 106 Bacteroides sp. CAG:633 synthesis unknown 107 Bacteroides common synthesis unknown 108 Helicobacter pylori synthesis Australia 109 Bacteroides sp. An51A synthesis unknown 110 Helicobacter sp. CLO-3 synthesis U.S. Table 2 : Summary of α-1,3-fucosyltransferases of the present invention SEQ ID NO organism source Country of Origin of Digital Serial Information 111 Helicobacter pylori synthesis Australia 112 Helicobacter pylori synthesis Australia 113 Basilea psittacipulmonis in psittacosis synthesis Switzerland 114 Azospirillum oryzae A2P synthesis Japan 115 Pyrromonas catarrhalis synthesis U.S. 116 Pyrromonas species synthesis U.K. 117 Selenomonas infelix synthesis unknown 118 Azospirillum lipoferum B510 synthesis Japan 119 Azospira sp. TSH64 synthesis Japan 120 Azospira sp. TSH100 synthesis Japan 121 Azospirillum brasilense synthesis Belgium 122 Azospira sp. B510 synthesis Japan 123 Planctopirus limnophila synthesis Germany 124 Pedobacter glucosidilyticus synthesis Germany 125 Vibrio butyricum TB synthesis unknown 126 Pyrromonas catarrhalis synthesis U.S. 127 Vibrio cellolyticus butyricum synthesis unknown 128 Rhizobium straminoryzae synthesis unknown 129 Lewinella xylanilytica synthesis South Korea 130 Homo sapiens synthesis unknown 131 Bisgaard Taxon 44 str. B96_3 synthesis Denmark 132 Methylovirgula ligni synthesis Netherlands Table 3 : Summary of other SEQ ID NOs described in the present invention SEQ ID NO organism source Country of Origin of Digital Serial Information 01 Escherichia coli W synthesis U.S. 02 Z. mobilis synthesis U.K. 03 Bifidobacterium adolescentis synthesis Germany 04 Escherichia coli K-12 MG1655 synthesis U.S. 05 Escherichia coli K-12 MG1655 synthesis U.S. 06 Escherichia coli K-12 MG1655 synthesis U.S. 07 Escherichia coli K-12 MG1655 synthesis U.S. 08 Escherichia coli K-12 MG1655 synthesis U.S. 09 Escherichia coli K-12 MG1655 synthesis U.S. 10 Escherichia coli K-12 MG1655 synthesis U.S. 11 Bacteroides fragilis synthesis U.K. 12 Escherichia coli K-12 MG1655 synthesis U.S. 13 Neisseria meningitidis MC58 synthesis U.K. 14 Escherichia coli K-12 MG1655 synthesis U.S. 15 Kluyveromyces lactis synthesis U.S. Culture conditions

Pre-incubation of the 96-well microtiter plate experiments started in frozen vials in 150 µL LB and incubated overnight at 37°C on an orbital shaker at 800 rpm. This culture was used as an inoculum in a 96-well square microtiter plate by diluting 400-fold with 400 µL of minimal medium. These final 96-well plates were then incubated at 37°C on an orbital shaker at 800 rpm for 72 h or less or longer. To measure the sugar concentration at the end of the culture experiment, samples of the whole culture fluid (= average of intracellular and extracellular sugar concentrations) were obtained from each well by boiling the culture fluid at 60°C for 15 min, after which the cells were briefly centrifuged .

Bioreactor pre-cultivation begins with an entire 1 mL frozen vial of a strain, inoculated in 250 mL or 500 mL of minimal medium in a 1 L or 2.5 L shake flask, and inoculated on an orbital shaker at 37°C. Incubate at 200 rpm for 24 hours. A 5 L bioreactor (250 mL inoculum in 2 L batch medium) was then inoculated; the process was controlled by MFCS control software (Sartorius Stedim Biotech, Melsungen, Germany). Cultivation conditions were set at 37°C and maximum agitation; pressure gas flow rates were strain and bioreactor dependent. The pH was controlled at 6.8 using 0.5 M H2S04 and 20% NH4OH. Cool the exhaust gas. When foaming occurs during fermentation, add 10% polysiloxane defoamer solution. optical density

The cell density of the cultures was usually monitored by measuring the optical density at 600 nm (Implen Nanophotometer NP80, Westburg, Belgium, or with a Spark 10M microplate reader, Tecan, Switzerland). Analytical Analysis

Standards such as (but not limited to) sucrose, lactose, 2'FL, 3-FL and DiFL were purchased from Carbosynth (UK), Elicityl (France) and IsoSep (Sweden).

Glycans were analyzed on a Waters Acquity H-stage UPLC with Evaporative Light Scattering Detector (ELSD) or refractive index (RI) detection. A volume of 0.7 µL was injected on a Waters Acquity UPLC BEH Amide column (2.1 × 100 mm; 130 Å; 1.7 µm) and an Acquity UPLC BEH Amide VanGuard column (130 Å, 2.1 × 5 mm). The column temperature was 50°C. The mobile phase consisted of a solution of 1/4 water and 2/4 acetonitrile to which 0.2% triethylamine was added. The method is isocratic at a flow rate of 0.130 mL/min. The drift tube temperature of the ELS detector was 50°C, and the N2 gas pressure was 50 psi, the gain was 200, and the data rate was 10 pps. The temperature of the RI detector was set to 35°C.

Glycans were also analyzed on a Waters Acquity H-stage UPLC with Refractive Index (RI) detection. A volume of 0.5 µL was injected on a Waters Acquity UPLC BEH Amide column (2.1 x 100 mm; 130 Å; 1.7 µm). The column temperature was 50°C. The mobile phase consisted of a mixture of 72% acetonitrile and 28% ammonium acetate buffer (100 mM) to which 0.1% triethylamine was added. The method is isocratic at a flow rate of 0.260 mL/min. The temperature of the RI detector was set to 35°C. Example 2. Materials and methods of Saccharomyces cerevisiae

Grow strains on synthetic defined yeast medium with complete supplement mix (SD CSM) or CSM drop-out (SD CSM-Ura, SD CSM-Trp, SD CSM-His) containing 6.7 g/L of Yeast nitrogen base without amino acids (YNB without AA, Difco), 20 g/L agar (Difco) (solid culture), 22 g/L glucose monohydrate or 20 g/L lactose and 0.79 g/L CSM or 0.77 g/L CSM-Ura, 0.77 g/L CSM-Trp or 0.77 g/L CSM-His (MP Biomedicals). Lactose, 2'FL and/or 3-FL were added as acceptors. strain

Saccharomyces cerevisiae BY4742 produced by Brachmann et al. (Yeast (1998) 14:115-32), available from Euroscarf culture collections, was used. All mutant strains were generated by homologous recombination or plastid transformation using the Gietz method (Yeast 11:355-360, 1995). plastid

In one example of production of GDP-fucose, yeast expressed the plastid p2a_2µ_Fuc (Chan 2013, Plasmid 70, 2-17) for the expression of foreign genes in Saccharomyces cerevisiae. This plastid contains an ampicillin resistance gene and a bacterial origin of replication to allow selection and maintenance in E. coli, and a 2µ yeast ori and Ura3 selectable marker for selection and maintenance in yeast. This plastid additionally contains a continuous transcription unit for a lactose permease such as, for example, LAC12 from Kluyveromyces lactis with SEQ ID NO 15, such as, for example, gmd from E. coli with SEQ ID NO 08 GDP-mannose 4,6-dehydratase and GDP-L-fucose synthase such as eg fcl from E. coli with SEQ ID NO 09. The yeast expression plastid p2a_2µ_Fuc2 was used as a surrogate expression plastid for the p2a_2µ_Fuc plastid containing a persistent transcription unit next to the ampicillin resistance gene, bacterial ori, 2µ yeast ori and the Ura3 selectable marker for: As e.g. lactose permease from LAC12 of Kluyveromyces lactis with SEQ ID NO 15, fucose permease as e.g. from fucP of Escherichia coli with SEQ ID NO 10 and as e.g. Bifunctional fucosokinase/fucose-1-phosphate guanosyltransferase of fkp of Bacteroides fragilis. For further production of 2'FL, 3-FL and DiFL, p2a_2µ_Fuc and its variant p2a_2µ_Fuc2 additionally contain at least one α-1,2-fucosyltransferase and at least one α-1,3-fucosyl Continuous transcription units of transferases, wherein the fucosyltransferases are selected from SEQ ID NOs 16 to 132 as shown in Tables 1 and 2.

Preferably, but not necessarily, fucosyltransferases, proteins involved in nucleotide-activated sugar synthesis, and/or membrane proteins are N-terminally and/or C-terminally fused to a SUMOstar tag (eg, obtained from pYSUMOstar, Life Sensors, Malvern, PA). ) to enhance its solubility.

Optionally, mutant yeast strains are modified with gene body insertion of persistent transcription units encoding companion proteins such as, for example, Hsp31, Hsp32, Hsp33, Sno4, Kar2, Ssb1, Sse1, Sse2, Ssa1, Ssa2, Ssa3, Ssa4 , Ssb2, Ecm10, Ssc1, Ssq1, Ssz1, Lhs1, Hsp82, Hsc82, Hsp78, Hsp104, Tcp1, Cct4, Cct8, Cct2, Cct3, Cct5, Cct6 or Cct7 (Gong et al., 2009, Mol. Syst. Biol. 5 : 275).

Plastids were maintained in host E. coli DH5α (F ⁻ , phi80d lacZ deltaM15, delta( lacZYA − argF ) U169, deoR , recA1 , endA1 , hsdR17(rk ⁻ , mk ⁺ ), phoA , supE44 , lambda ⁻ purchased from Invitrogen , thi -1, gyrA96 , rel A1). Heterologous and Homologous Expression

Genes to be expressed, whether from plastids or gene bodies, are synthesized synthetically by one of the following companies: DNA2.0, Gen9, IDT or Twist Bioscience. Expression can be further facilitated by optimizing codon usage to that of the expression host. Genetic optimization using vendor tools. Culture conditions

Generally, yeast strains are initially grown on SD CSM disks to obtain single colonies. The plates were grown at 30°C for 2-3 days. Starting with a single colony, the preculture was grown overnight at 30 °C in 5 mL with shaking at 200 rpm. Subsequent 125 mL shake flask experiments were seeded with 2% of this preculture in 25 mL of medium. The flasks were incubated at 30°C with orbital shaking at 200 rpm. gene expression promoter

The gene was expressed using a synthetic persistent promoter, as described by Blazeck (Biotechnology and Bioengineering, Vol. 109, No. 11, 2012). Example 3. Production of 2'FL , 3 -containing E. coli hosts using modified E. coli hosts expressing alpha -1,2- fucosyltransferases and alpha - 1,3 -fucosyltransferases from compatibilities - Oligosaccharide mixture of FL and DiFL

E. coli K12 strains modified for GDP-fucose production as described in Example 1 were sequentially expressed for alpha-1,2-fucosyltransferases selected from SEQ ID NOs 17 and 18 Transformation of a first plastid of a persistent expression construct and a second compatible plastid expressing a persistent expression construct selected from the group consisting of α-1,3-fucosyltransferases of SEQ ID NOs 111 to 117 . Table 4 presents an overview of novel strains with transcription units for expressing the respective fucosyltransferases. The novel strains were evaluated in growth experiments according to the culture conditions provided in Example 1, wherein the medium contained sucrose and lactose. Each strain was grown in four biological replicates in 96-well plates. After 72 h of incubation, the broth was collected and the sugar mixture was analyzed on UPLC. The relative yields of 2'FL, 3-FL and DiFL obtained in the culture fluid samples of each strain as provided in Table 5 were obtained from the mixture of strains (ie 2'FL, 3-FL and DiFL) The average production titer of each fucosylated sugar was calculated by dividing by the sum of the average production titers of all fucosylated sugars. It can be concluded from Table 5 that all new strains expressing one alpha-1,2-fucosyltransferase and one alpha-1,3-fucosyltransferase produced 2'FL, 3-FL and A mixture of DiFL. Table 5 also shows that the ratio of the oligosaccharides 2'FL, 3-FL and DiFL produced in the mixture of novel strains can be used to express each fucosyl by selecting the fucosyltransferases combined in the strains and selecting The transcription unit of the transferase is precisely regulated. Table 4 : Use in modified E. coli strains to express an α-1,2-fucosyltransferase (2FT) and an α-1,3-fucosyltransferase (3FT) from different plastids transcription unit strain Fucosyl-transferase transcription unit Promoter UTR SEQ ID NO of 2FT or 3FT terminator F1 2FT MutalikP10 ThrA_BCD2 17 T7early 3FT MutalikP6 OmpC_BCD5 113 T7early F2 2FT MutalikP5 GalE_IptFG 17 T7early 3FT MutalikP6 OmpC_BCD5 113 T7early F3 2FT apFAB142 OmpC_BCD18 17 T7early 3FT MutalikP6 OmpC_BCD5 113 T7early F4 2FT MutalikP12 Gene10-LeuAB-BCD2 18 rnpB_T1 3FT MutalikP6 OmpC_BCD5 113 T7early F5 2FT MutalikP5 GalE_IptFG 17 T7early 3FT MutalikP5 Gene10_LeuL 114 rnpB_T1 F6 2FT apFAB142 OmpC_BCD18 17 T7early 3FT MutalikP5 Gene10_LeuL 114 rnpB_T1 F7 2FT MutalikP12 Gene10-LeuAB-BCD2 18 rnpB_T1 3FT MutalikP5 Gene10_LeuL 114 rnpB_T1 F8 2FT MutalikP5 Gene10_LeuL 18 L3S2P53 3FT P14 p14-UTR 111 T7early F9 2FT MutalikP5 Gene10_LeuL 18 L3S2P53 3FT P14 p14-UTR 112 rnpB_T1 F10 2FT MutalikP5 Gene10_LeuL 18 L3S2P53 3FT P14 Gene10-LeuAB-BCD2 113 T7early F11 2FT MutalikP5 Gene10_LeuL 18 L3S2P53 3FT P14 Gene10-LeuAB-BCD2 115 T7early F12 2FT MutalikP5 Gene10_LeuL 18 L3S2P53 3FT P14 Gene10-LeuAB-BCD2 116 T7early F13 2FT MutalikP5 Gene10_LeuL 18 L3S2P53 3FT P14 Gene10-LeuAB-BCD2 117 T7early Table 5 : Single α-1,2-fucosyltransferase and one α-1,3-fucosyltransferase according to Table 3 when grown in minimal medium with sucrose and lactose Relative yield of 2'FL, 3-FL and DiFL in the case of modified E. coli strains (%) strain 2 ' FL ( % ) 3FL ( % ) DiFL ( % ) F1 27 ± 0.4 30 ± 0.2 43 ± 0.4 F2 23 ± 0.2 37 ± 0.4 40 ± 0.2 F3 20 ± 0.2 44 ± 0.4 36 ± 0.2 F4 22 ± 0.3 39 ± 0.2 39 ± 0.3 F5 31 ± 0.4 17 ± 0.5 52 ± 0.4 F6 27 ± 0.2 23 ± 0.8 50 ± 0.2 F7 37 ± 0.1 4.0 ± 0.2 59 ± 0.1 F8 48 ± 1.6 19 ± 0.6 33 ± 1.6 F9 33 ± 1.2 27 ± 0.2 40 ± 1.2 F10 15 ± 0.3 51 ± 1.1 34 ± 0.3 F11 16 ± 1.1 45 ± 1.2 39 ± 1.1 F12 12 ± 0.5 61 ± 6.8 27 ± 0.5 F13 3.0 ± 5.0 57 ± 3.9 40 ± 5.0 Example 4. Production of 2'FL , 3- _ _ _ _ _ Oligosaccharide mixture of FL and DiFL

E. coli strains modified for GDP-fucose production as described in Example 1 were further transformed with an expression plastid with two persistent transcription units, one for alpha-1,2 - Fucosyltransferase and another for alpha-1,3-fucosyltransferase. Several expressing plastids containing different combinations of α-1,2-fucosyltransferase and α-1,3-fucosyltransferase were cloned as described in Example 1. Table 6 presents a summary of novel strains with transcription units for expressing the respective fucosyltransferases. The novel strains were evaluated in growth experiments according to the culture conditions provided in Example 1, wherein the medium contained sucrose and lactose. Each strain was grown in four biological replicates in 96-well plates. After 72 h of incubation, the broth was collected and the sugar mixture was analyzed on UPLC.

The relative yields of 2'FL, 3-FL and DiFL obtained in the broth samples of each strain as provided in Table 7 were obtained from the mixture of strains (ie 2'FL, 3-FL and DiFL) The average production titer of each fucosylated sugar was calculated by dividing by the sum of the average production titers of all fucosylated sugars. It can be concluded from Table 7 that all new strains expressing one α-1,2-fucosyltransferase and one α-1,3-fucosyltransferase from a single plastid produced 2'FL, A mixture of 3-FL and DiFL. Table 7 also shows that the ratios of the oligosaccharides 2'FL, 3-FL and DiFL produced in the mixture of novel strains can be used to express each fucosyl by selecting the fucosyltransferases combined in the strains and selecting The transcription unit of the transferase is precisely regulated. Table 6 : Transcription units used in modified E. coli strains to express an alpha-1,2-fucosyltransferase and an alpha-1,3-fucosyltransferase from a plastid strain Fucosyl - transferase transcription unit Promoter UTR SEQ ID NO of 2FT or 3FT terminator F14 2FT MutalikP9 Gene10_LeuL 16 L3S2P53 3FT MutalikP6 OmpC_BCD5 113 rnpB_T1 F15 2FT MutalikP5 Gene10_LeuL 16 L3S2P53 3FT MutalikP6 OmpC_BCD5 113 rnpB_T1 F16 2FT apFAB146 Gene10_LeuL 16 L3S2P53 3FT MutalikP6 OmpC_BCD5 113 rnpB_T1 F17 2FT MutalikP9 Gene10_LeuL 17 L3S2P53 3FT MutalikP6 OmpC_BCD5 114 rnpB_T1 F18 2FT MutalikP5 Gene10_LeuL 17 L3S2P53 3FT MutalikP6 OmpC_BCD5 114 rnpB_T1 F19 2FT apFAB146 Gene10_LeuL 17 L3S2P53 3FT MutalikP6 OmpC_BCD5 114 rnpB_T1 F20 2FT MutalikP9 Gene10_LeuL 18 L3S2P53 3FT MutalikP6 OmpC_BCD5 114 rnpB_T1 F21 2FT MutalikP5 Gene10_LeuL 18 L3S2P53 3FT MutalikP6 OmpC_BCD5 114 rnpB_T1 F22 2FT apFAB146 Gene10_LeuL 18 L3S2P53 3FT MutalikP6 OmpC_BCD5 114 rnpB_T1 F23 2FT MutalikP9 Gene10_LeuL 19 L3S2P53 3FT MutalikP6 OmpC_BCD5 114 rnpB_T1 F24 2FT MutalikP9 Gene10_LeuL 16 L3S2P53 3FT MutalikP10 OmpC_BCD5 113 rnpB_T1 F25 2FT MutalikP5 Gene10_LeuL 16 L3S2P53 3FT MutalikP10 OmpC_BCD5 113 rnpB_T1 F26 2FT apFAB146 Gene10_LeuL 16 L3S2P53 3FT MutalikP10 OmpC_BCD5 113 rnpB_T1 F27 2FT MutalikP9 Gene10_LeuL 17 L3S2P53 3FT MutalikP10 OmpC_BCD5 114 rnpB_T1 F28 2FT MutalikP5 Gene10_LeuL 17 L3S2P53 3FT MutalikP10 OmpC_BCD5 114 rnpB_T1 F29 2FT apFAB146 Gene10_LeuL 17 L3S2P53 3FT MutalikP10 OmpC_BCD5 114 rnpB_T1 F30 2FT MutalikP9 Gene10_LeuL 18 L3S2P53 3FT MutalikP10 OmpC_BCD5 114 rnpB_T1 F31 2FT MutalikP5 Gene10_LeuL 18 L3S2P53 3FT MutalikP10 OmpC_BCD5 114 rnpB_T1 F32 2FT apFAB146 Gene10_LeuL 18 L3S2P53 3FT MutalikP10 OmpC_BCD5 114 rnpB_T1 F33 2FT MutalikP9 Gene10_LeuL 19 L3S2P53 3FT MutalikP10 OmpC_BCD5 114 rnpB_T1 F34 2FT MutalikP5 Gene10_LeuL 19 L3S2P53 3FT MutalikP10 OmpC_BCD5 114 rnpB_T1 Table 7 : Single α-1,2-fucosyltransferase and one α-1,3-fucosyltransferase according to Table 5 when grown in minimal medium with sucrose and lactose Relative yield of 2'FL, 3-FL and DiFL in the case of modified E. coli strains (%) strain 2 ' FL ( % ) 3FL ( % ) DiFL ( % ) F14 1.0 ± 0.1 29 ± 0.4 70 ± 0.1 F15 9.0 ± 0.2 4 ± 0.1 87 ± 0.2 F16 21 ± 0.3 2.0 ± 0.1 77 ± 0.3 F17 1.0 ± 0 85 ± 0.5 14 ± 0 F18 3.0 ± 0.1 76 ± 0.5 21 ± 0.1 F19 9.0 ± 0.4 58 ± 0.9 33 ± 0.4 F20 3.0 ± 0 62 ± 1.6 35 ± 0 F21 10 ± 0.1 40 ± 0.4 50 ± 0.1 F22 16 ± 0.3 30 ± 0.3 54 ± 0.3 F23 11 ± 0.5 4.0 ± 0.2 85 ± 0.5 F24 1.0 ± 0 33 ± 1.1 66 ± 0 F25 6.0 ± 0.2 5.0 ± 0.3 89 ± 0.2 F26 13 ± 0.1 3.0 ± 0.1 84 ± 0.1 F27 1.0 ± 0 92 ± 0.2 7.0 ± 0 F28 1.0 ± 0 86 ± 0.4 13 ± 0 F29 4.0 ± 0.3 73 ± 1.4 23 ± 0.3 F30 2.0 ± 0.1 83 ± 0.2 15 ± 0.1 F31 6.0 ± 0.1 65 ± 0.6 29 ± 0.1 F32 9.0 ± 0.2 58 ± 0.4 33 ± 0.2 F33 11 ± 0.1 12 ± 0.5 77 ± 0.1 F34 15 ± 2.8 12 ± 6.1 73 ± 2.8 Example 5. Production of an oligosaccharide mixture comprising 2'FL , 3-FL and DiFL using a modified E. coli host

E. coli strains modified for GDP-fucose production as described in Example 1 were further adapted for intracellular lactose synthesis by gene body deletion of the lacZ , glk and galETKM operons and for use in Gene body insertion of the IgtB of Neisseria meningitidis with SEQ ID NO 13 and the persistent transcription unit from E. coli with SEQ ID NO 14 of UDP-glucose 4-epimerase (galE). In the next step, mutant E. coli strains were transformed with expression plastids with two persistent transcription units for expression of H. pylori alpha-1,2-fucoid with SEQ ID NO 16 Glycosyltransferase and Helicobacter pylori alpha-1,3-fucosyltransferase having SEQ ID NO 111. When evaluated in growth experiments according to the culture conditions provided in Example 1, the novel strain produced a mixture of oligosaccharides comprising 2'FL, 3-FL and DiFL in a whole broth sample, wherein the medium contained sucrose as a carbon source and No precursors. Example 6. Production of an oligosaccharide mixture comprising at least 80% DiFL using a modified E. coli host

E. coli strains modified for GDP-fucose production as described in Example 1 were further transformed with an expression plastid with two persistent transcription units, one for alpha-1,2 - Fucosyltransferase and another for alpha-1,3-fucosyltransferase. Several expressing plastids containing different combinations of α-1,2-fucosyltransferase and α-1,3-fucosyltransferase were cloned as described in Example 1. Table 8 presents a summary of novel strains with transcription units for expressing the respective fucosyltransferases. The novel strains were evaluated in growth experiments according to the culture conditions provided in Example 1, wherein the medium contained sucrose and lactose. Each strain was grown in four biological replicates in 96-well plates. After 72 h of incubation, the broth was collected and the sugar mixture was analyzed on UPLC.

The relative yields of 2'FL, 3-FL and DiFL obtained in the culture fluid samples of each strain as provided in Table 9 were obtained from those produced in the mixture of strains (ie 2'FL, 3-FL and DiFL) The average production titer of each fucosylated sugar was calculated by dividing by the sum of the average production titers of all fucosylated sugars. From Table 9 it can be concluded that all new strains produced mixtures containing more than 80% DiFL. Table 9 also shows that these strains produced less than 6% 3-FL. Table 8 : Transcription units used in modified E. coli strains to express an alpha-1,2-fucosyltransferase and an alpha-1,3-fucosyltransferase from a plastid strain Fucosyl - transferase transcription unit Promoter UTR SEQ ID NO of 2FT or 3FT terminator F15 2FT MutalikP5 Gene10_LeuL 16 L3S2P53 3FT MutalikP6 OmpC_BCD5 113 rnpB_T1 F23 2FT MutalikP9 Gene10_LeuL 19 L3S2P53 3FT MutalikP6 OmpC_BCD5 114 rnpB_T1 F37 2FT MutalikP5 Gene10_LeuL 16 L3S2P53 3FT MutalikP10 OmpC_BCD5 113 rnpB_T1 F38 2FT apFAB146 Gene10_LeuL 16 L3S2P53 3FT MutalikP10 OmpC_BCD5 113 rnpB_T1 Table 9 : Performance of one alpha-1,2-fucosyltransferase and one alpha-1,3-fucosyltransferase according to Table 7 when grown in minimal medium with sucrose and lactose Relative yield of 2'FL, 3-FL and DiFL in the case of modified E. coli strains (%) strain 2 ' FL ( % ) 3FL ( % ) DiFL ( % ) F15 9.0 ± 0.2 4 ± 0.1 87 ± 0.2 F23 11 ± 0.5 4.0 ± 0.2 85 ± 0.5 F37 6.0 ± 0.2 5.0 ± 0.3 89 ± 0.2 F38 13 ± 0.1 3.0 ± 0.1 84 ± 0.1 Example 7. Production of Oligosaccharide Mixtures Comprising 2'FL , 3-FL and DiFL Using Modified E. coli Hosts in Fed-Batch Fermentation

Mutant E. coli strains as described in Examples 3 and 4 were further evaluated in a fed-batch fermentation process. Fed-batch fermentation was performed as described in Example 1 at the bioreactor scale. In these examples, sucrose was used as the carbon source and lactose was added to the batch medium as the acceptor. Regular broth samples were obtained and UPLC as described in Example 1 was used to measure the production of 2'FL, 3-FL and DiFL. The experiments demonstrated that the broth samples obtained throughout the fed-batch phase contained oligosaccharide mixtures of 2'FL, 3-FL and DiFL (Table 10). Since the ratio of lactose, 2'FL, 3-FL and DiFL changes over time during fed-batch, it can be done during the fermentation process by interrupting the fermentation at desired times during the fed-batch stage process to control. The final broth of fermentation runs DiFLF004 to DiFLF008 achieved a DiFL purity of >80% or >90% as measured by the total amount of 2'FL, 3-FL, DiFL and lactose in the mixture. Table 10 : Whole broth samples of modified E. coli strains at various time points during a fed-batch fermentation operation with sucrose as carbon source and lactose as precursor added to the batch medium 2'FL, 3-FL and DiFL were produced in relative percentages (%). The modified strains tested included strains F4 and F7 from Example 3 in runs DiFLF003 and DiFLF004, respectively, and strains F23, F26, F19, and F21 from Example 4 in runs DiFLF005, DiFLF006, DiFLF007, and DiFLF008, respectively . ID time point 2 ' FL ( % ) 3-FL ( % ) DiFL ( % ) Lactose ( % ) DiFLF003 1 0 0 0 100 DiFLF003 2 5.0 12 3.7 79 DiFLF003 3 twenty one 51 28 0 DiFLF003 4 6.7 48 45 0 DiFLF004 1 0 0 0 100 DiFLF004 2 14 1.0 8.8 76 DiFLF004 3 66 3.0 31 0 DiFLF004 4 2.2 0.5 97 0 DiFLF005 1 0 0 0 100 DiFLF005 2 14 0.3 14 72 DiFLF005 3 48 1.5 35 15 DiFLF005 4 15 0.1 85 0 DiFLF006 1 0 0 0 100 DiFLF006 2 6.0 0 17 77 DiFLF006 3 11 6.1 63 20 DiFLF006 4 9.0 10 80 0.6 DiFLF007 1 0 0 0 100 DiFLF007 2 11 4.0 8.1 77 DiFLF007 3 18 twenty three 59 0 DiFLF007 4 0.1 9.2 91 0 DiFLF008 1 0 0 0 100 DiFLF008 2 8.6 5.6 7.3 79 DiFLF008 3 36 9.8 54 0 DiFLF008 4 0.1 3.4 97 0 Example 8. Production of 2'FL , 3 - FL and _ _ _ DiFL Oligosaccharide Mixture

The E. coli strain modified for GDP-fucose production as described in Example 1 was further modified with gene body insertion of a persistent transcription unit for H. pylori a1 with SEQ ID NO 16 , 2-fucosyltransferase HpFutC. In the next step, this novel strain, as well as the parental strain that produces GDP-fucose and lacks HpFutC, contains two enzymes for al,2-fucosyltransferase and al,3-fucosyltransferase Expression of persistent transcription units by plastid transformation. Therefore, strains were generated to express one or two al,2-fucosyltransferases and one al,3-fucosyltransferase. A summary of the strains and their respective fucosyltransferase expression modules is provided in Table 11. The novel strains were evaluated in growth experiments according to the culture conditions provided in Example 1, wherein the medium contained sucrose and lactose. Each strain was grown in four biological replicates in 96-well plates. After 72 h of incubation, the broth was collected and the sugar mixture was analyzed on UPLC. The relative yields of 2'FL, 3-FL and DiFL obtained in the broth samples of each strain as provided in Table 12 were obtained from the mixture of strains (ie 2'FL, 3-FL and DiFL) The average production titer of each fucosylated sugar was calculated by dividing by the sum of the average production titers of all fucosylated sugars. From Table 12 it can be concluded that compared to the strains expressing one al,2-fucosyltransferase and one al,3-fucosyltransferase, the strains from H. pylori with SEQ ID NO 16 exhibited Novel strains with additional a1,2-fucosyltransferase and one selected a1,2-fucosyltransferase and one selected a1,3-fucosyltransferase produced similar amounts of 2'FL, lower Amounts of 3-FL and higher amounts of DiFL. Table 11 : Transcription units used in modified E. coli strains to express one or two alpha-1,2-fucosyltransferases and one alpha-1,3-fucosyltransferase strain Fucosyl - transferase transcription unit Promoter UTR SEQ ID NO of 2FT or 3FT terminator F39 2FT apFAB146 Gene10_LeuL 17 L3S2P53 3FT MutalikP6 OmpC_BCD5 114 rnpB_T1 F40 2FT P14 p14-UTR 16 L3S2P53 2FT apFAB146 Gene10_LeuL 17 L3S2P53 3FT MutalikP6 OmpC_BCD5 114 rnpB_T1 F41 2FT MutalikP5 Gene10_LeuL 18 L3S2P53 3FT MutalikP6 OmpC_BCD5 114 rnpB_T1 F42 2FT P14 p14-UTR 16 L3S2P53 2FT MutalikP5 Gene10_LeuL 18 L3S2P53 3FT MutalikP6 OmpC_BCD5 114 rnpB_T1 Table 12 : Performance when grown in minimal medium with sucrose and lactose according to Table 10 one or two alpha-1,2-fucosyltransferases and one alpha-1,3-fucosyltransferase Relative yield (%) of 2'FL, 3-FL and DiFL with a single modified E. coli strain of enzymes strain 2 ' FL ( % ) 3FL ( % ) DiFL ( % ) F39 12 37 51 F40 13 16 71 F41 9 33 58 F42 9 twenty two 69 Example 9. Production of an oligosaccharide mixture comprising 2'FL , 3-FL and DiFL

Another example provides the use of al,2-fucosyltransferases and al,3-fucosyltransferases having SEQ ID NOs 16 to 110 and SEQ ID NOs 111 to 132, respectively, of the present invention. Such enzymes are produced in cell-free expression systems such as, but not limited to, the PURExpress system (NEB) or in host organisms such as, but not limited to, E. coli or Saccharomyces cerevisiae, after which the enzymes listed above can be Isolated and optionally further purified.

A combination of al,2-fucosyltransferase and al,3-fucosyltransferase selected from the above enzyme extracts or purified enzymes is combined with GDP-fucose and a buffer component such as Tris-HCl or HEPES and lactose) were added to the reaction mixture together as acceptors. The reaction mixture is then incubated at a certain temperature (eg 37°C) for a certain amount of time (eg 24 hours) during which lactose will be converted enzymatically using GDP-fucose to 2'fucosyllactose, 3' Fucosyllactose and 2',3-difucosyllactose.

Alternatively, a single a1,2-fucosyltransferase or a1,3-fucosyltransferase extract or purified enzyme, respectively, can be added to the same reaction mixture, but containing 3-FL or 2'FL as acceptor . The reaction mixture is then incubated at a certain temperature (eg, 37°C) for a certain amount of time (eg, 24 hours), during which 3-FL or 2'FL is converted to 2',3 by enzymes using GDP-fucose - Difucosyllactose.

The 2'FL, 3-FL and/or DiFL are then isolated from the reaction mixture by methods known in the art. If preferred, further purification of 2'FL and/or 3-FL and/or diFL can be performed. At the end of the reaction or after isolation and/or purification, the production of 2'FL, 3-FL and DiFL was measured as described in Example 1. Example 10. Production of an oligosaccharide mixture comprising 2'FL , 3-FL and DiFL using a modified S. cerevisiae host

Saccharomyces cerevisiae strains were adapted for GDP-fucose production with an α-1,2-fucosyltransferase and an α-1 with yeast expressing plastids (variants of p2a_2µ_Fuc) as described in Example 2 , Expression of 3-fucosyltransferase, the expression plastid comprises a continuous transcription unit for the following: lactose permease (LAC12) from Kluyveromyces lactis with SEQ ID NO 15, from lactose permease with SEQ ID NO 15 GDP-mannose 4,6-dehydratase (gmd) from Escherichia coli with ID NO 8, GDP-L-fucose synthase (fcl) from Escherichia coli with SEQ ID NO 9, from Escherichia coli with SEQ ID NO 16 α-1,2-fucosyltransferase from Helicobacter pylori and α-1,3-fucosyltransferase from Helicobacter pylori with SEQ ID NO 111. The mutant yeast strain produced a mixture of oligosaccharides containing 2'FL, 3-FL and DiFL when evaluated on SD CSM-Ura omission medium containing lactose as acceptor. Example 11. Production of an oligosaccharide mixture comprising 2'FL , 3-FL and DiFL using a modified E. coli host

In the next example, the mutant E. coli strains described in Examples 3, 4, 5, and 6 were further modified with gene body insertion for a persistent transcription unit for a membrane protein selected from a list comprising : MdfA from S. mogeni (UniProt ID A0A2T7ANQ9), MdfA from Citrobacter johnsonii (UniProt ID D4BC23), MdfA from Escherichia coli (UniProt ID P0AEY8), from Yorkia reginsburg MdfA (UniProt ID G9Z5F4), iceT from E. coli (UniProt ID A0A024L207) or iceT from Citrobacter japonicus (UniProt ID D4B8A6). The novel strains were evaluated in growth experiments according to the culture conditions provided in Example 1, wherein the medium contained sucrose and lactose, for the production of oligosaccharide mixtures comprising 2'FL, 3-FL and DiFL. Example 12. Materials and Methods of Bacillus subtilis Medium

Two different media were used, namely enriched Luria broth (LB) and minimal medium for shake flasks (MMsf). Minimal medium uses a trace element mixture.

The trace element mixture consisted of 0.735 g/L CaCl2.2H2O, 0.1 g/L MnCl2.2H2O, 0.033 g/L CuCl2.2H2O, 0.06 g/L CoCl2.6H2O, 0.17 g/L ZnCl2, 0.0311 g/L H3BO4, 0.4 It is composed of g/L Na2EDTA.2H2O and 0.06 g/L Na2MoO4. The ferric citrate solution contained 0.135 g/L FeCl3.6H2O, 1 g/L sodium citrate (Hoch 1973 PMC1212887).

Luria Broth (LB) medium was composed of 1% pancreas (Difco, Erembodegem, Belgium), 0.5% yeast extract (Difco) and 0.5% sodium chloride ( VWR, Leuven, Belgium). Luria Broth agar (LBA) plates consisted of LB medium supplemented with 12 g/L agar (Difco, Erembodegem, Belgium).

Minimal medium for shake flask (MMsf) experiments containing 2.00 g/L (NH4)2SO4, 7.5 g/L KH2PO4, 17.5 g/L K2HPO4, 1.25 g/L sodium citrate, 0.25 g/L MgSO4.7H2O, 0.05 g/L L tryptophan, 10 to 30 g/L glucose or another carbon source (including (but not limited to) fructose, maltose, sucrose, glycerol and maltotriose as specified in the examples), 10 ml/L trace Elemental mixture and 10 ml/L ferric citrate solution. The medium was set to pH 7 with 1 M KOH. Depending on the experiment, lactose, 2'FL and/or 3-FL can be added as acceptors.

Sterilize complex media (eg LB) by autoclaving (121°C, 21') and sterilize minimal media by filtration (0.22 µm Sartorius). When necessary, the medium was made selective by adding antibiotics such as zeocin (20 mg/L). Strains, plastids and mutations

Bacillus subtilis 168, available from the Bacillus Gene Reserve (Ohio, USA).

Genetically deleted plastids via Cre/lox were constructed as described by Yan et al. (Appl. & Environm. Microbial., September 2008, pp. 5556-5562). Gene disruption occurs via homologous recombination with linear DNA and transformation via electroporation, as described by Xue et al. (J. Microb. Meth. 34 (1999) 183-191). The method of gene knockout is described by Liu et al. (Metab. Engine. 24 (2014) 61-69). This method uses 1000 bp of homology upstream and downstream of the gene of interest.

Integration vectors as described by Popp et al. (Sci. Rep., 2017, 7, 15158) are used as expression vectors and can be further used for genome integration if necessary. Promoters suitable for expression can be derived from partial repositories (iGems): sequence id: Bba_K143012, Bba_K823000, Bba_K823002 or Bba_K823003. Colonization can be performed using Gibson Assembly, Golden Gate assembly, Cliva assembly, LCR or restriction ligation.

In one embodiment of the production of lactose-based oligosaccharides, mutant strains of B. subtilis are generated to contain a gene encoding a lactose importer (such as, for example, E. coli lacY having SEQ ID NO 12). For 2'FL, 3FL and diFL production, alpha-1,2-fucosyltransferases selected from the list comprising SEQ ID NOs 16 to 110 and alpha-1,2-fucosyltransferases selected from the list comprising SEQ ID NOs 111 to 132 will be used An expression construct for alpha-1,3-fucosyltransferase was additionally added to the strain. Heterologous and Homologous Expression

Genes to be expressed, whether from plastids or gene bodies, are synthesized synthetically by one of the following companies: DNA2.0, Gen9, Twist Biosciences or IDT.

Expression can be further facilitated by optimizing codon usage to that of the expression host. Genetic optimization using vendor tools. Culture conditions

Pre-incubation of 96-well microtiter plate experiments was performed starting from a frozen vial or from a single colony in an LB plate, in 150 µL of LB, and incubated overnight at 37°C on an orbital shaker at 800 rpm. This culture was used as an inoculum in a 96-well square microtiter plate by diluting 400-fold with 400 µL of MMsf medium. Each strain was grown in multiple wells of a 96-well plate as biological replicates. These final 96-well plates were then incubated at 37°C on an orbital shaker at 800 rpm for 72 h or less or longer. At the end of the culture experiment, samples were taken from each well to measure the supernatant concentration (extracellular sugar concentration, after briefly centrifuging cells for 5 min), or by boiling the culture at 90°C for 15 min before briefly centrifuging cells Or measure by boiling at 60°C for 60 min (= concentration of whole culture medium, concentration of intracellular and extracellular sugars, as defined herein).

In addition, cultures were diluted to measure optical density at 600 nm. The Cell Potency Index or CPI was determined by dividing the oligosaccharide concentration by the biomass (as a relative percentage compared to the reference strain). It is empirically determined that biomass is approximately 1/3 the optical density measured at 600 nm. Example 13. Production of an oligosaccharide mixture comprising 2'FL , 3-FL and DiFL using a modified Bacillus subtilis host

Bacillus subtilis strains were modified as described in Example 12 by gene body insertion of persistent transcription units for lactose permease (LacY) from E. coli having SEQ ID NO 12 and a selective From the alpha-1,2-fucosyltransferases comprising the listing of SEQ ID NOs 16-110 and an alpha-1,3-fucosyltransferase selected from the listings comprising SEQ ID NOs 111-132. Novel strains were evaluated for production of 2'FL, 3-FL and DiFL in growth experiments on MMsf medium containing lactose according to the culture conditions provided in Example 12. After 72 h of incubation, the broth was collected and analyzed for sugars on UPLC. Example 14. Materials and methods of Corynebacterium glutamicum culture medium

Two different media were used, namely enriched tryptone-yeast (TY) medium and minimal medium for shake flask (MMsf). Minimal medium uses 1000x stock trace element mix.

The trace element mixture was composed of 10 g/L CaCl2, 10 g/L FeSO4.7H2O, 10 g/L MnSO4.H2O, 1 g/L ZnSO4.7H2O, 0.2 g/L CuSO4, 0.02 g/L NiCl2.6H2O, 0.2 g/L biotin (pH 7.0) and 0.03 g/L protocatechuic acid.

Minimal medium for shake flask (MMsf) experiments containing 20 g/L (NH4)2SO4, 5 g/L urea, 1 g/L KH2PO4, 1 g/L K2HPO4, 0.25 g/L MgSO4.7H2O, 42 g/L MOPS , 10 to 30 g/L glucose or another carbon source (including (but not limited to) fructose, maltose, sucrose, glycerol and maltotriose as specified in the examples) and 1 ml/L trace element mixture. Depending on the experiment, lactose, 2'FL and/or 3-FL were added as acceptors.

TY medium was composed of 1.6% pancreas (Difco, Ellen Bodheim, Belgium), 1% yeast extract (Difco) and 0.5% sodium chloride (VWR, Leuven, Belgium). The TY agar (TYA) plate system consisted of TY medium supplemented with 12 g/L agar (Difco, Ellen Bodheim, Belgium).

Sterilize complex media (eg TY) by autoclaving (121°C, 21') and sterilize minimal media by filtration (0.22 µm Sartorius). When necessary, the medium is made selective by the addition of antibiotics (eg, kanamycin, ampicillin). Strains and Mutations

Corynebacterium glutamicum ATCC 13032 is available from the American Type Culture Collection.

Integrating plastid vectors based on Cre/loxP technology as described by Suzuki et al. (Appl. Microbiol. Biotechnol., April 2005, 67(2):225-33) and as described by Okibe et al. (Journal of The temperature-sensitive shuttle vectors described in Microbiological Methods 85, 2011, 155-163) were constructed for gene deletion, mutation and insertion. Suitable promoters for (heterologous) gene expression can be derived from Yim et al. (Biotechnol. Bioeng., 2013 Nov, 110(11):2959-69). Colonization can be performed using Gibson assembly, Golden Gate assembly, Cliva assembly, LCR or restriction ligation.

In one embodiment of the production of lactose-based oligosaccharides, a mutant strain of Corynebacterium glutamicum is produced to contain a gene encoding a lactose importer (such as, for example, E. coli lacY having SEQ ID NO 12). For 2'FL, 3FL and diFL production, alpha-1,2-fucosyltransferases selected from the list comprising SEQ ID NOs 16 to 110 and alpha-1,2-fucosyltransferases selected from the list comprising SEQ ID NOs 111 to 132 will be used An expression construct for alpha-1,3-fucosyltransferase was additionally added to the strain. Heterologous and Homologous Expression

Genes to be expressed, whether from plastids or gene bodies, are synthesized synthetically by one of the following companies: DNA2.0, Gen9, Twist Biosciences or IDT.

Expression can be further facilitated by optimizing codon usage to that of the expression host. Genetic optimization using vendor tools. Culture conditions

Pre-incubation of 96-well microtiter plate experiments was performed starting from a frozen vial or from a single colony in a TY plate, in 150 µL of TY, and incubated overnight at 37°C on an orbital shaker at 800 rpm. This culture was used as an inoculum in a 96-well square microtiter plate by diluting 400-fold with 400 µL of MMsf medium. Each strain was grown in multiple wells of a 96-well plate as biological replicates. These final 96-well plates were then incubated at 37°C on an orbital shaker at 800 rpm for 72 h or less or longer. At the end of the incubation experiment, samples were taken from each well to measure the supernatant concentration (extracellular sugar concentration, after briefly centrifuging cells for 5 min), or by boiling the culture at 60°C for 15 min before briefly centrifuging cells Measurements (= concentration of whole broth, concentration of intracellular and extracellular sugars, as defined herein).

In addition, cultures were diluted to measure optical density at 600 nm. The Cell Potency Index or CPI was determined by dividing the measured oligosaccharide concentration in the whole broth by the biomass (as a relative percentage compared to the reference strain). It is empirically determined that biomass is approximately 1/3 the optical density measured at 600 nm. Example 15. Production of an oligosaccharide mixture comprising 2'FL , 3-FL and DiFL using a modified Corynebacterium glutamicum host

Corynebacterium glutamicum strains were modified by gene body insertion of persistent transcription units for lactose permease (LacY) from Escherichia coli with SEQ ID NO 12 as described in Example 14 , sucrose permease CscB from Escherichia coli W with SEQ ID NO 01, fructokinase Frk from Zymomonas mobilis with SEQ ID NO 02, sucrose from Bifidobacterium adolescentis with SEQ ID NO 03 Phosphorylase BaSP and one alpha-1,2-fucosyltransferase selected from the list comprising SEQ ID NOs 16 to 110 and one alpha-1,3- selected from the list comprising SEQ ID NOs 111 to 132 Fucosyltransferase. Novel strains were evaluated for the production of 2'FL, 3-FL and DiFL in growth experiments on MMsf medium containing lactose according to the culture conditions provided in Example 14. After 72 h of incubation, the broth was collected and analyzed for sugars on UPLC. Example 16. Materials and Methods of Chlamydomonas reinhardtii Medium

Chlamydomonas reinhardtii cells were cultured in Tris-acetate-phosphate (TAP) medium (pH 7.0). TAP medium used 1000X stock Hutner's trace element mix. Hertner's trace element mixture consisted of 50 g/L Na2EDTA.H2O (Titriplex III), 22 g/L ZnSO4.7H2O, 11.4 g/L H3BO3, 5 g/L MnCl2.4H2O, 5 g/L FeSO4.7H2O , 1.6 g/L CoCl2.6H2O, 1.6 g/L CuSO4.5H2O and 1.1 g/L (NH4)6MoO3.

TAP medium contains 2.42 g/L Tris (para(hydroxymethyl)aminomethane), 25 mg/L salt stock solution, 0.108 g/L K2HPO4, 0.054 g/L KH2PO4, and 1.0 mL/L glacial acetic acid. The salt stock solution consisted of 15 g/L NH4CL, 4 g/L MgSO4.7H2O, and 2 g/L CaCl2.2H2O. As precursors for sugar synthesis, precursors such as, for example, galactose, glucose, fructose, fucose can be added. Lactose, 2'FL and/or 3-FL can be added as acceptors. The medium was sterilized by autoclaving (121°C, 21'). For stock cultures on slanted agar, use TAP medium containing 1% agar (with purified high strength, 1000 g/cm2). Strains, plastids and mutations

Chlamydomonas reinhardtii wild type strain 21gr (CC-1690, wild type, mt+), 6145C (CC-1691, wild type, mt-), CC-125 (137c, wild type, mt+), CC-124 (137c, wild type) Type, mt-) can be purchased from the University of Minnesota (University of Minnesota, U.S.A.) Chlamydomonas Resource Center (Chlamydomonas Resource Center) (https://www.chlamycollection.org).

Expression plastids were derived from pSI103, eg, available from the Chlamydomonas Resource Center. Colonization can be performed using Gibson assembly, Golden Gate assembly, Cliva assembly, LCR or restriction ligation. Suitable promoters for (heterologous) gene expression can be derived, for example, from Scranton et al. (Algal Res. 2016, 15: 135-142). Targeted genetic modification, such as gene knockout or gene replacement, can be performed using the Crispr-Cas technology as described, for example, by Jiang et al. (Eukaryotic Cell 2014, 13(11): 1465-1469).

Transformation via electroporation was performed as described by Wang et al. (Biosci. Rep. 2019, 39: BSR2018210). Cells were grown in liquid TAP medium under constant aeration and continuous light at a light intensity of 8000 Lx until the cell density reached 1.0-2.0 x 107 cells/ml. Next, cells were seeded into freshly prepared liquid TAP medium at a concentration of 1.0×10 6 cells/ml and grown under continuous light for 18-20 h until the cell density reached 4.0×10 6 cells/ml. Next, cells were harvested by centrifugation at 1250 g for 5 min at room temperature, washed and resuspended with pre-cooled liquid TAP medium (Sigma, USA) containing 60 mM sorbitol and frozen for 10 min. Next, 250 µL of the cell suspension (corresponding to 5.0 x 107 cells) was placed in a pre-chilled 0.4 cm electroporation cuvette (400 ng/mL) with 100 ng of plastid DNA. Electroporation was performed using a BTX ECM830 electroporation device (1575 Ω, 50 μFD) with 6 pulses of 500 V each with a pulse length of 4 ms and a pulse interval of 100 ms. Immediately after electroporation, the cuvette was placed on ice for 10 min. Finally, the cell suspension was transferred to a 50 ml conical centrifuge tube containing 10 mL of freshly prepared liquid TAP medium with 60 mM sorbitol and allowed to recover overnight in the dark with slow shaking. After overnight recovery, cells were re-harvested and seeded by starch embedding onto selective 1.5% (w/v) agar-TAP dishes containing either ampicillin (100 mg/L) or chloramphenicol (100 mg/v). L). The disks were then incubated at 23+-0.5°C under continuous illumination with a light intensity of 8000 Lx. Cells were analyzed after 5-7 days.

In one embodiment of the production of UDP-galactose, C. reinhardtii cells are modified with transcriptional units comprising genes encoding galactokinase from Arabidopsis (KIN, UniProt ID Q9SEE5) and from A. thaliana ) of UDP-sugar pyrophosphorylase (USP) (UniProt ID Q9C5I1).

In one example of producing GDP-fucose, C. reinhardtii cells were modified with a transcription unit for GDP-fucose synthase from Arabidopsis (GER1, UniProt ID 049213).

In one example of fucosylation of lactose, C. reinhardtii cells may be modified with expression plastids comprising transcription units for alpha-1,2 selected from the list comprising SEQ ID NOs 16-110 - Fucosyltransferases and alpha-1,3-fucosyltransferases selected from the list comprising SEQ ID NOs 111 to 132. Heterologous and Homologous Expression

Genes to be expressed, whether from plastids or gene bodies, are synthesized synthetically by one of the following companies: DNA2.0, Gen9, Twist Biosciences or IDT.

Expression can be further facilitated by optimizing codon usage to that of the expression host. Genetic optimization using vendor tools. Culture conditions

Chlamydomonas reinhardtii cells were cultured in selective TAP-agar dishes at 23 +/- 0.5°C with a light intensity of 8000 Lx on a 14/10 h light/dark cycle. Cells were analyzed after 5 to 7 days in culture.

For high density cultures, cells can be grown in a closed system as described by Chen et al. (Bioresour. Technol. 2011, 102: 71-81) and Johnson et al. (Biotechnol. Prog. 2018, 34: 811-827), Such as for example vertical or horizontal tube photobioreactors, stirred tank photobioreactors or flat plate photobioreactors. Example 17. Production of an oligosaccharide mixture comprising 2'FL , 3-FL and DiFL in mutant Chlamydomonas reinhardtii cells

Chlamydomonas reinhardtii cells were engineered for the production of UDP-Gal as described in Example 16 with gene body insertion of a persistent transcription unit comprising encoding galactose kinase (KIN, UniProt ID Q9SEE5) and Arabidopsis gene for UDP-sugar pyrophosphorylase (USP) (UniProt ID Q9C5I1). In the next step, cells are modified with gene body insertion of persistent transcription units comprising GDP-fucose synthase from Arabidopsis (GER1, UniProt ID 049213) and a protein selected from the group consisting of SEQ ID NO. Alpha-1 ,2-fucosyltransferases of the listings 16 to 110 and an alpha-1 ,3-fucosyltransferases selected from the listings comprising SEQ ID NOs 111 to 132. According to the culture conditions provided in Example 16, novel strains were evaluated in growth experiments on TAP agar plates containing galactose as a precursor. After 5 days of incubation, cells were harvested and analyzed on UPLC for 2'FL, 3-FL and DiFL production. Example 18. Materials and Methods for Animal Cells Isolation of Mesenchymal Stem Cells from Adipose Tissue of Different Mammals

Fresh adipose tissue is obtained from slaughterhouses (eg, cattle, pigs, sheep, chickens, ducks, catfish, snakes, frogs) or from liposuction (eg, in the case of humans, after signed informed consent) and maintained at in phosphate buffered saline supplemented with antibiotics. Enzymatic digestion of adipose tissue followed by centrifugation to isolate mesenchymal stem cells. The isolated interleaf stem cells are transferred to cell culture flasks and grown under standard growth conditions (eg, 37°C, 5% CO2). Initial media included DMEM-F12, RPMI and Alpha-MEM media (supplemented with 15% fetal bovine serum) and 1% antibiotics. Then after the first passage, the medium was replaced with medium supplemented with 10% fetal bovine serum (FBS). For example, Ahmad and Shakoori (2013, Stem Cell Regen Med. 9(2): 29-36), herein incorporated by reference in its entirety for all purposes, describe the methods described in this example some variations of it. Isolation of mesenchymal stem cells from milk

This example illustrates the isolation of mesenchymal stem cells from milk collected under sterile conditions from a human or any other mammal, such as those described herein. An equal volume of phosphate buffered saline was added to the diluted milk followed by centrifugation for 20 min. Cell pellets were washed three times with phosphate buffered saline, and cells were seeded in cell culture flasks in DMEM-F12, RPMI and Alpha-MEM medium supplemented with 10% fetal bovine serum and 1% antibiotics under standard culture conditions. For example, Hassiotou et al. (2012, Stem Cells. 30(10): 2164-2174), which is incorporated herein by reference in its entirety for all purposes, describe some of the methods described in this example. some variations. Differentiate stem cells using 2D and 3D culture systems

The isolated mesenchymal cells can be differentiated into mammary-like epithelial cells and luminal cells in 2D and 3D culture systems. See eg, Huynh et al. 1991. Exp Cell Res. 197(2): 191-199; Gibson et al. 1991, In Vitro Cell Dev Biol Anim. 27(7): 585-594; Blatchford et al. 1999; Animal Cell Technology': Basic & Applied Aspects, Springer, Dordrecht. 141-145; Williams et al. 2009, Breast Cancer Res 11(3): 26-43; and Arevalo et al. 2015, Am J Physiol Cell Physiol. 310( 5): C348-C356; each of which is incorporated herein by reference in its entirety for all purposes.

For 2D cultures, isolated cells were initially seeded in culture dishes in growth medium supplemented with 10 ng/ml epithelial growth factor and 5 pg/ml insulin. At confluence, cells were fed for 48 h with growth medium supplemented with 2% fetal bovine serum, 1% penicillin-streptomycin (100 U/ml penicillin, 100 ug/ml streptomycin) and 5 pg/ml insulin. To induce differentiation, cells were fed complete growth medium containing 5 pg/ml insulin, 1 pg/ml cortisol, 0.65 ng/ml triiodothyronine, 100 nM dexamethasone and 1 pg/ml prolactin. After 24 h, serum was removed from the complete induction medium.

For 3D culture, isolated cells were trypsinized and cultured in Matrigel, hyaluronic acid, or ultra-low attachment surface culture dishes for six days, and incubated by adding epithelial growth factor supplemented with 10 ng/ml and 5 pg/ml insulin. Growth medium to induce differentiation and lactation. At confluence, cells were fed for 48 h with growth medium supplemented with 2% fetal bovine serum, 1% penicillin-streptomycin (100 U/ml penicillin, 100 ug/ml streptomycin) and 5 pg/ml insulin. To induce differentiation, cells were fed complete growth medium containing 5 pg/ml insulin, 1 pg/ml cortisol, 0.65 ng/ml triiodothyronine, 100 nM dexamethasone and 1 pg/ml prolactin. After 24 h, serum was removed from the complete induction medium. Method of making mammary gland-like cells

Mammalian cells were induced to pluripotency by reprogramming with viral vectors encoding Oct4, Sox2, Klf4 and c-Myc. The resulting reprogrammed cells were then cultured in Mammocult medium (obtained from Stem Cell Technologies) or breast cell enrichment medium (DMEM, 3% FBS, estrogen, progesterone, heparin, corticosterone, insulin, EGF) to This is transformed into mammary gland-like cells from which the expression of selected milk components can be induced. Alternatively, epigenetic remodeling is performed using a remodeling system such as CRISPR/Cas9 to activate selected genes of interest, such as casein, alpha-lactalbumin to be persisted, to allow expression of their respective proteins, and/or Selected endogenous genes are down-regulated and/or knocked out, as described, for example, in WO21067641, which is incorporated herein by reference in its entirety for all purposes. nourish

Complete growth medium included high glucose DMEM/F12, 10% FBS, 1% NEAA, 1% pen/strep, 1% ITS-X, 1% F-Glu, 10 ng/ml EGF, and 5 pg/ml hydrocortisone. Complete lactation medium including high glucose DMEM/F12, 1% NEAA, 1% pen/strep, 1% ITS-X, 1% F-Glu, 10 ng/ml EGF, 5 pg/ml hydrocortisone, and 1 pg/ml Prolactin (5ug/ml in Hyunh 1991). Cells were seeded on collagen-coated flasks in complete growth medium at a density of 20,000 cells/cm 2 and left to adhere and expand in complete growth medium for 48 hours, after which the medium was changed to Complete lactation medium. After exposure to lactation medium, cells begin to differentiate and stop growing. Within about a week, cells begin to secrete lactation products, such as milk lipids, lactose, casein, and whey, into the medium. The desired concentration of lactation medium can be achieved by concentration or dilution by ultrafiltration. The desired salt balance of the lactation medium can be achieved by dialysis, eg, to remove undesired metabolites from the medium. The hormones and other growth factors used can be purified by resins, such as selective extraction using nickel resins to remove His-tagged growth factors, to further reduce the level of contaminants in the lactated product. Example 19. Production of an oligosaccharide mixture comprising 2'FL , 3-FL and DiFL in non-mammary adult stem cells

Isolated mesenchymal cells and reprogrammed into mammary gland-like cells as described in Example 18 were modified by CRISPR-CAS to overexpress GDP-fucose synthase GFUS (UniProt ID Q13630) from Homo sapiens, a species selected from An alpha-1,2-fucosyltransferase comprising the listing of SEQ ID NOs 16-110 and an alpha-1,3-fucosyltransferase selected from the listing comprising SEQ ID NOs 111-132. Cells were seeded on collagen-coated flasks in complete growth medium at a density of 20,000 cells/cm 2 and left to adhere and expand in complete growth medium for 48 hours, after which the medium was changed to Complete lactation medium for about 7 days. After culturing as described in Example 18, cells were subjected to UPLC to analyze the production of an oligosaccharide mixture comprising 2'FL, 3-FL and DiFL. Example 20. Production of an oligosaccharide mixture comprising 2'FL , 3-FL and DiFL using a modified E. coli host

In the next example, the mutant E. coli strains described in Examples 3, 4, 5, 6 and 11 were evaluated in growth experiments according to the culture conditions provided in Example 1, wherein the culture medium contained sucrose and lactose, and additionally supplied 2'FL and/or 3-FL to the medium for the production of oligosaccharide mixtures comprising 2'FL, 3-FL and DiFL.

none

none

                                  
           <![CDATA[ <110> Inbiose N.V., Belgium]]>
           <![CDATA[ <120> Production of fucosylated lactose structures by cells]]>
           <![CDATA[ <130> 024-TW]]>
           <![CDATA[ <140>TW 110129388]]>
           <![CDATA[ <141> 2021-08-10]]>
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           <![CDATA[ <151> 2020-08-10]]>
           <![CDATA[ <150> EP 20190204.6]]>
           <![CDATA[ <151> 2020-08-10]]>
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          Met Ala Leu Asn Ile Pro Phe Arg Asn Ala Tyr Tyr Arg Phe Ala Ser
          1 5 10 15
          Ser Tyr Ser Phe Leu Phe Phe Ile Ser Trp Ser Leu Trp Trp Ser Leu
                      20 25 30
          Tyr Ala Ile Trp Leu Lys Gly His Leu Gly Leu Thr Gly Thr Glu Leu
                  35 40 45
          Gly Thr Leu Tyr Ser Val Asn Gln Phe Thr Ser Ile Leu Phe Met Met
              50 55 60
          Phe Tyr Gly Ile Val Gln Asp Lys Leu Gly Leu Lys Lys Pro Leu Ile
          65 70 75 80
          Trp Cys Met Ser Phe Ile Leu Val Leu Thr Gly Pro Phe Met Ile Tyr
                          85 90 95
          Val Tyr Glu Pro Leu Leu Gln Ser Asn Phe Ser Val Gly Leu Ile Leu
                      100 105 110
          Gly Ala Leu Phe Phe Gly Leu Gly Tyr Leu Ala Gly Cys Gly Leu Leu
                  115 120 125
          Asp Ser Phe Thr Glu Lys Met Ala Arg Asn Phe His Phe Glu Tyr Gly
              130 135 140
          Thr Ala Arg Ala Trp Gly Ser Phe Gly Tyr Ala Ile Gly Ala Phe Phe
          145 150 155 160
          Ala Gly Ile Phe Phe Ser Ile Ser Pro His Ile Asn Phe Trp Leu Val
                          165 170 175
          Ser Leu Phe Gly Ala Val Phe Met Met Ile Asn Met Arg Phe Lys Asp
                      180 185 190
          Lys Asp His Gln Cys Val Ala Ala Asp Ala Gly Gly Val Lys Lys Glu
                  195 200 205
          Asp Phe Ile Ala Val Phe Lys Asp Arg Asn Phe Trp Val Phe Val Ile
              210 215 220
          Phe Ile Val Gly Thr Trp Ser Phe Tyr Asn Ile Phe Asp Gln Gln Leu
          225 230 235 240
          Phe Pro Val Phe Tyr Ser Gly Leu Phe Glu Ser His Asp Val Gly Thr
                          245 250 255
          Arg Leu Tyr Gly Tyr Leu Asn Ser Phe Gln Val Val Leu Glu Ala Leu
                      260 265 270
          Cys Met Ala Ile Ile Pro Phe Phe Val Asn Arg Val Gly Pro Lys Asn
                  275 280 285
          Ala Leu Leu Ile Gly Val Val Ile Met Ala Leu Arg Ile Leu Ser Cys
              290 295 300
          Ala Leu Phe Val Asn Pro Trp Ile Ile Ser Leu Val Lys Leu Leu His
          305 310 315 320
          Ala Ile Glu Val Pro Leu Cys Val Ile Ser Val Phe Lys Tyr Ser Val
                          325 330 335
          Ala Asn Phe Asp Lys Arg Leu Ser Ser Thr Ile Phe Leu Ile Gly Phe
                      340 345 350
          Gln Ile Ala Ser Ser Leu Gly Ile Val Leu Leu Ser Thr Pro Thr Gly
                  355 360 365
          Ile Leu Phe Asp His Ala Gly Tyr Gln Thr Val Phe Phe Ala Ile Ser
              370 375 380
          Gly Ile Val Cys Leu Met Leu Leu Phe Gly Ile Phe Phe Leu Ser Lys
          385 390 395 400
          Lys Arg Glu Gln Ile Val Met Glu Thr Pro Val Pro Ser Ala Ile
                          405 410 415
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          Met Lys Asn Asp Lys Lys Ile Tyr Gly Cys Ile Glu Gly Gly Gly Thr
          1 5 10 15
          Lys Phe Met Leu Ala Leu Ile Asp Ser Asp Arg Lys Met Leu Ala Val
                      20 25 30
          Glu Arg Val Pro Thr Thr Thr Pro Glu Glu Thr Leu Gly Lys Ser Val
                  35 40 45
          Glu Phe Phe Lys Lys Ala Leu Pro Gln Tyr Ala Asp Ser Phe Ala Ser
              50 55 60
          Phe Gly Ile Ala Ser Phe Gly Pro Leu Cys Leu Asp Arg Lys Ser Pro
          65 70 75 80
          Lys Trp Gly Tyr Ile Thr Asn Thr Pro Lys Pro Phe Trp Pro Asn Thr
                          85 90 95
          Asp Val Val Thr Pro Phe Lys Glu Ala Phe Gly Cys Pro Val Glu Ile
                      100 105 110
          Asp Thr Asp Val Asn Gly Ala Ala Leu Ala Glu Asn Phe Trp Gly Ala
                  115 120 125
          Ser Lys Gly Thr His Thr Ser Val Tyr Val Thr Val Gly Thr Gly Phe
              130 135 140
          Gly Gly Gly Val Leu Ile Asp Gly Lys Pro Ile His Gly Leu Ala His
          145 150 155 160
          Pro Glu Met Gly His Gly Ile Pro Ile Arg His Pro Asp Asp Arg Asp
                          165 170 175
          Phe Glu Gly Cys Cys Pro Tyr His Gly Gly Cys Tyr Glu Gly Leu Ala
                      180 185 190
          Ser Gly Thr Ala Ile Arg Lys Arg Trp Gly Lys Ala Leu Asn Glu Met
                  195 200 205
          Glu Pro Ala Glu Phe Glu Lys Ala Arg Glu Ile Ile Ala Phe Tyr Leu
              210 215 220
          Ala His Phe Asn Val Thr Leu Gln Ala Phe Ile Ser Pro Glu Arg Ile
          225 230 235 240
          Val Phe Gly Gly Gly Val Met His Val Asp Gly Met Leu Ala Ser Val
                          245 250 255
          Arg Arg Gln Thr Ala Glu Ile Ala Asn Ser Tyr Phe Glu Gly Ala Asp
                      260 265 270
          Phe Glu Lys Ile Ile Val Leu Pro Gly Leu Gly Asp Gln Ala Gly Met
                  275 280 285
          Met Gly Ala Phe Ala Leu Ala Leu Ala Ala Glu Asn Lys
              290 295 300
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          Met Lys Asn Lys Val Gln Leu Ile Thr Tyr Ala Asp Arg Leu Gly Asp
          1 5 10 15
          Gly Thr Ile Lys Ser Met Thr Asp Ile Leu Arg Thr Arg Phe Asp Gly
                      20 25 30
          Val Tyr Asp Gly Val His Ile Leu Pro Phe Phe Thr Pro Phe Asp Gly
                  35 40 45
          Ala Asp Ala Gly Phe Asp Pro Ile Asp His Thr Lys Val Asp Glu Arg
              50 55 60
          Leu Gly Ser Trp Asp Asp Val Ala Glu Leu Ser Lys Thr His Asn Ile
          65 70 75 80
          Met Val Asp Ala Ile Val Asn His Met Ser Trp Glu Ser Lys Gln Phe
                          85 90 95
          Gln Asp Val Leu Ala Lys Gly Glu Glu Ser Glu Tyr Tyr Pro Met Phe
                      100 105 110
          Leu Thr Met Ser Ser Val Phe Pro Asn Gly Ala Thr Glu Glu Asp Leu
                  115 120 125
          Ala Gly Ile Tyr Arg Pro Arg Pro Gly Leu Pro Phe Thr His Tyr Lys
              130 135 140
          Phe Ala Gly Lys Thr Arg Leu Val Trp Val Ser Phe Thr Pro Gln Gln
          145 150 155 160
          Val Asp Ile Asp Thr Asp Ser Asp Lys Gly Trp Glu Tyr Leu Met Ser
                          165 170 175
          Ile Phe Asp Gln Met Ala Ala Ser His Val Ser Tyr Ile Arg Leu Asp
                      180 185 190
          Ala Val Gly Tyr Gly Ala Lys Glu Ala Gly Thr Ser Cys Phe Met Thr
                  195 200 205
          Pro Lys Thr Phe Lys Leu Ile Ser Arg Leu Arg Glu Glu Gly Val Lys
              210 215 220
          Arg Gly Leu Glu Ile Leu Ile Glu Val His Ser Tyr Tyr Lys Lys Gln
          225 230 235 240
          Val Glu Ile Ala Ser Lys Val Asp Arg Val Tyr Asp Phe Ala Leu Pro
                          245 250 255
          Pro Leu Leu Leu His Ala Leu Ser Thr Gly His Val Glu Pro Val Ala
                      260 265 270
          His Trp Thr Asp Ile Arg Pro Asn Asn Ala Val Thr Val Leu Asp Thr
                  275 280 285
          His Asp Gly Ile Gly Val Ile Asp Ile Gly Ser Asp Gln Leu Asp Arg
              290 295 300
          Ser Leu Lys Gly Leu Val Pro Asp Glu Asp Val Asp Asn Leu Val Asn
          305 310 315 320
          Thr Ile His Ala Asn Thr His Gly Glu Ser Gln Ala Ala Thr Gly Ala
                          325 330 335
          Ala Ala Ser Asn Leu Asp Leu Tyr Gln Val Asn Ser Thr Tyr Tyr Ser
                      340 345 350
          Ala Leu Gly Cys Asn Asp Gln His Tyr Ile Ala Ala Arg Ala Val Gln
                  355 360 365
          Phe Phe Leu Pro Gly Val Pro Gln Val Tyr Tyr Val Gly Ala Leu Ala
              370 375 380
          Gly Lys Asn Asp Met Glu Leu Leu Arg Lys Thr Asn Asn Gly Arg Asp
          385 390 395 400
          Ile Asn Arg His Tyr Tyr Ser Thr Ala Glu Ile Asp Glu Asn Leu Lys
                          405 410 415
          Arg Pro Val Val Lys Ala Leu Asn Ala Leu Ala Lys Phe Arg Asn Glu
                      420 425 430
          Leu Asp Ala Phe Asp Gly Thr Phe Ser Tyr Thr Thr Asp Asp Asp Thr
                  435 440 445
          Ser Ile Ser Phe Thr Trp Arg Gly Glu Thr Ser Gln Ala Thr Leu Thr
              450 455 460
          Phe Glu Pro Lys Arg Gly Leu Gly Val Asp Asn Thr Thr Pro Val Ala
          465 470 475 480
          Met Leu Glu Trp Glu Asp Ser Ala Gly Asp His Arg Ser Asp Asp Leu
                          485 490 495
          Ile Ala Asn Pro Pro Val Val Ala
                      500
           <![CDATA[ <210> 4]]>
           <![CDATA[ <211> 264]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Escherichia coli K-12 MG1655]]>
           <![CDATA[ <400> 4]]>
          Met Lys Gln Tyr Leu Glu Leu Met Gln Lys Val Leu Asp Glu Gly Thr
          1 5 10 15
          Gln Lys Asn Asp Arg Thr Gly Thr Gly Thr Leu Ser Ile Phe Gly His
                      20 25 30
          Gln Met Arg Phe Asn Leu Gln Asp Gly Phe Pro Leu Val Thr Thr Lys
                  35 40 45
          Arg Cys His Leu Arg Ser Ile Ile His Glu Leu Leu Trp Phe Leu Gln
              50 55 60
          Gly Asp Thr Asn Ile Ala Tyr Leu His Glu Asn Asn Val Thr Ile Trp
          65 70 75 80
          Asp Glu Trp Ala Asp Glu Asn Gly Asp Leu Gly Pro Val Tyr Gly Lys
                          85 90 95
          Gln Trp Arg Ala Trp Pro Thr Pro Asp Gly Arg His Ile Asp Gln Ile
                      100 105 110
          Thr Thr Val Leu Asn Gln Leu Lys Asn Asp Pro Asp Ser Arg Arg Ile
                  115 120 125
          Ile Val Ser Ala Trp Asn Val Gly Glu Leu Asp Lys Met Ala Leu Ala
              130 135 140
          Pro Cys His Ala Phe Phe Gln Phe Tyr Val Ala Asp Gly Lys Leu Ser
          145 150 155 160
          Cys Gln Leu Tyr Gln Arg Ser Cys Asp Val Phe Leu Gly Leu Pro Phe
                          165 170 175
          Asn Ile Ala Ser Tyr Ala Leu Leu Val His Met Met Ala Gln Gln Cys
                      180 185 190
          Asp Leu Glu Val Gly Asp Phe Val Trp Thr Gly Gly Asp Thr His Leu
                  195 200 205
          Tyr Ser Asn His Met Asp Gln Thr His Leu Gln Leu Ser Arg Glu Pro
              210 215 220
          Arg Pro Leu Pro Lys Leu Ile Ile Lys Arg Lys Pro Glu Ser Ile Phe
          225 230 235 240
          Asp Tyr Arg Phe Glu Asp Phe Glu Ile Glu Gly Tyr Asp Pro His Pro
                          245 250 255
          Gly Ile Lys Ala Pro Val Ala Ile
                      260
           <![CDATA[ <210> 5]]>
           <![CDATA[ <211> 391]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Escherichia coli K-12 MG1655]]>
           <![CDATA[ <400> 5]]>
          Met Gln Lys Leu Ile Asn Ser Val Gln Asn Tyr Ala Trp Gly Ser Lys
          1 5 10 15
          Thr Ala Leu Thr Glu Leu Tyr Gly Met Glu Asn Pro Ser Ser Gln Pro
                      20 25 30
          Met Ala Glu Leu Trp Met Gly Ala His Pro Lys Ser Ser Ser Arg Val
                  35 40 45
          Gln Asn Ala Ala Gly Asp Ile Val Ser Leu Arg Asp Val Ile Glu Ser
              50 55 60
          Asp Lys Ser Thr Leu Leu Gly Glu Ala Val Ala Lys Arg Phe Gly Glu
          65 70 75 80
          Leu Pro Phe Leu Phe Lys Val Leu Cys Ala Ala Gln Pro Leu Ser Ile
                          85 90 95
          Gln Val His Pro Asn Lys His Asn Ser Glu Ile Gly Phe Ala Lys Glu
                      100 105 110
          Asn Ala Ala Gly Ile Pro Met Asp Ala Ala Glu Arg Asn Tyr Lys Asp
                  115 120 125
          Pro Asn His Lys Pro Glu Leu Val Phe Ala Leu Thr Pro Phe Leu Ala
              130 135 140
          Met Asn Ala Phe Arg Glu Phe Ser Glu Ile Val Ser Leu Leu Gln Pro
          145 150 155 160
          Val Ala Gly Ala His Pro Ala Ile Ala His Phe Leu Gln Gln Pro Asp
                          165 170 175
          Ala Glu Arg Leu Ser Glu Leu Phe Ala Ser Leu Leu Asn Met Gln Gly
                      180 185 190
          Glu Glu Lys Ser Arg Ala Leu Ala Ile Leu Lys Ser Ala Leu Asp Ser
                  195 200 205
          Gln Gln Gly Glu Pro Trp Gln Thr Ile Arg Leu Ile Ser Glu Phe Tyr
              210 215 220
          Pro Glu Asp Ser Gly Leu Phe Ser Pro Leu Leu Leu Asn Val Val Lys
          225 230 235 240
          Leu Asn Pro Gly Glu Ala Met Phe Leu Phe Ala Glu Thr Pro His Ala
                          245 250 255
          Tyr Leu Gln Gly Val Ala Leu Glu Val Met Ala Asn Ser Asp Asn Val
                      260 265 270
          Leu Arg Ala Gly Leu Thr Pro Lys Tyr Ile Asp Ile Pro Glu Leu Val
                  275 280 285
          Ala Asn Val Lys Phe Glu Ala Lys Pro Ala Asn Gln Leu Leu Thr Gln
              290 295 300
          Pro Val Lys Gln Gly Ala Glu Leu Asp Phe Pro Ile Pro Val Asp Asp
          305 310 315 320
          Phe Ala Phe Ser Leu His Asp Leu Ser Asp Lys Glu Thr Thr Ile Ser
                          325 330 335
          Gln Gln Ser Ala Ala Ile Leu Phe Cys Val Glu Gly Asp Ala Thr Leu
                      340 345 350
          Trp Lys Gly Ser Gln Gln Leu Gln Leu Lys Pro Gly Glu Ser Ala Phe
                  355 360 365
          Ile Ala Ala Asn Glu Ser Pro Val Thr Val Lys Gly His Gly Arg Leu
              370 375 380
          Ala Arg Val Tyr Asn Lys Leu
          385 390
           <![CDATA[ <210> 6]]>
           <![CDATA[ <211> 456]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Escherichia coli K-12 MG1655]]>
           <![CDATA[ <400> 6]]>
          Met Lys Lys Leu Thr Cys Phe Lys Ala Tyr Asp Ile Arg Gly Lys Leu
          1 5 10 15
          Gly Glu Glu Leu Asn Glu Asp Ile Ala Trp Arg Ile Gly Arg Ala Tyr
                      20 25 30
          Gly Glu Phe Leu Lys Pro Lys Thr Ile Val Leu Gly Gly Asp Val Arg
                  35 40 45
          Leu Thr Ser Glu Thr Leu Lys Leu Ala Leu Ala Lys Gly Leu Gln Asp
              50 55 60
          Ala Gly Val Asp Val Leu Asp Ile Gly Met Ser Gly Thr Glu Glu Ile
          65 70 75 80
          Tyr Phe Ala Thr Phe His Leu Gly Val Asp Gly Gly Ile Glu Val Thr
                          85 90 95
          Ala Ser His Asn Pro Met Asp Tyr Asn Gly Met Lys Leu Val Arg Glu
                      100 105 110
          Gly Ala Arg Pro Ile Ser Gly Asp Thr Gly Leu Arg Asp Val Gln Arg
                  115 120 125
          Leu Ala Glu Ala Asn Asp Phe Pro Pro Val Asp Glu Thr Lys Arg Gly
              130 135 140
          Arg Tyr Gln Gln Ile Asn Leu Arg Asp Ala Tyr Val Asp His Leu Phe
          145 150 155 160
          Gly Tyr Ile Asn Val Lys Asn Leu Thr Pro Leu Lys Leu Val Ile Asn
                          165 170 175
          Ser Gly Asn Gly Ala Ala Gly Pro Val Val Asp Ala Ile Glu Ala Arg
                      180 185 190
          Phe Lys Ala Leu Gly Ala Pro Val Glu Leu Ile Lys Val His Asn Thr
                  195 200 205
          Pro Asp Gly Asn Phe Pro Asn Gly Ile Pro Asn Pro Leu Leu Pro Glu
              210 215 220
          Cys Arg Asp Asp Thr Arg Asn Ala Val Ile Lys His Gly Ala Asp Met
          225 230 235 240
          Gly Ile Ala Phe Asp Gly Asp Phe Asp Arg Cys Phe Leu Phe Asp Glu
                          245 250 255
          Lys Gly Gln Phe Ile Glu Gly Tyr Tyr Ile Val Gly Leu Leu Ala Glu
                      260 265 270
          Ala Phe Leu Glu Lys Asn Pro Gly Ala Lys Ile Ile His Asp Pro Arg
                  275 280 285
          Leu Ser Trp Asn Thr Val Asp Val Val Thr Ala Ala Gly Gly Thr Pro
              290 295 300
          Val Met Ser Lys Thr Gly His Ala Phe Ile Lys Glu Arg Met Arg Lys
          305 310 315 320
          Glu Asp Ala Ile Tyr Gly Gly Glu Met Ser Ala His His Tyr Phe Arg
                          325 330 335
          Asp Phe Ala Tyr Cys Asp Ser Gly Met Ile Pro Trp Leu Leu Val Ala
                      340 345 350
          Glu Leu Val Cys Leu Lys Asp Lys Thr Leu Gly Glu Leu Val Arg Asp
                  355 360 365
          Arg Met Ala Ala Phe Pro Ala Ser Gly Glu Ile Asn Ser Lys Leu Ala
              370 375 380
          Gln Pro Val Glu Ala Ile Asn Arg Val Glu Gln His Phe Ser Arg Glu
          385 390 395 400
          Ala Leu Ala Val Asp Arg Thr Asp Gly Ile Ser Met Thr Phe Ala Asp
                          405 410 415
          Trp Arg Phe Asn Leu Arg Thr Ser Asn Thr Glu Pro Val Val Arg Leu
                      420 425 430
          Asn Val Glu Ser Arg Gly Asp Val Pro Leu Met Glu Ala Arg Thr Arg
                  435 440 445
          Thr Leu Leu Thr Leu Leu Asn Glu
              450 455
           <![CDATA[ <210> 7]]>
           <![CDATA[ <211> 478]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Escherichia coli K-12 MG1655]]>
           <![CDATA[ <400> 7]]>
          Met Ala Gln Ser Lys Leu Tyr Pro Val Val Met Ala Gly Gly Ser Gly
          1 5 10 15
          Ser Arg Leu Trp Pro Leu Ser Arg Val Leu Tyr Pro Lys Gln Phe Leu
                      20 25 30
          Cys Leu Lys Gly Asp Leu Thr Met Leu Gln Thr Thr Ile Cys Arg Leu
                  35 40 45
          Asn Gly Val Glu Cys Glu Ser Pro Val Val Ile Cys Asn Glu Gln His
              50 55 60
          Arg Phe Ile Val Ala Glu Gln Leu Arg Gln Leu Asn Lys Leu Thr Glu
          65 70 75 80
          Asn Ile Ile Leu Glu Pro Ala Gly Arg Asn Thr Ala Pro Ala Ile Ala
                          85 90 95
          Leu Ala Ala Leu Ala Ala Lys Arg His Ser Pro Glu Ser Asp Pro Leu
                      100 105 110
          Met Leu Val Leu Ala Ala Asp His Val Ile Ala Asp Glu Asp Ala Phe
                  115 120 125
          Arg Ala Ala Val Arg Asn Ala Met Pro Tyr Ala Glu Ala Gly Lys Leu
              130 135 140
          Val Thr Phe Gly Ile Val Pro Asp Leu Pro Glu Thr Gly Tyr Gly Tyr
          145 150 155 160
          Ile Arg Arg Gly Glu Val Ser Ala Gly Glu Gln Asp Met Val Ala Phe
                          165 170 175
          Glu Val Ala Gln Phe Val Glu Lys Pro Asn Leu Glu Thr Ala Gln Ala
                      180 185 190
          Tyr Val Ala Ser Gly Glu Tyr Tyr Trp Asn Ser Gly Met Phe Leu Phe
                  195 200 205
          Arg Ala Gly Arg Tyr Leu Glu Glu Leu Lys Lys Tyr Arg Pro Asp Ile
              210 215 220
          Leu Asp Ala Cys Glu Lys Ala Met Ser Ala Val Asp Pro Asp Leu Asn
          225 230 235 240
          Phe Ile Arg Val Asp Glu Glu Ala Phe Leu Ala Cys Pro Glu Glu Ser
                          245 250 255
          Val Asp Tyr Ala Val Met Glu Arg Thr Ala Asp Ala Val Val Val Pro
                      260 265 270
          Met Asp Ala Gly Trp Ser Asp Val Gly Ser Trp Ser Ser Leu Trp Glu
                  275 280 285
          Ile Ser Ala His Thr Ala Glu Gly Asn Val Cys His Gly Asp Val Ile
              290 295 300
          Asn His Lys Thr Glu Asn Ser Tyr Val Tyr Ala Glu Ser Gly Leu Val
          305 310 315 320
          Thr Thr Val Gly Val Lys Asp Leu Val Val Val Gln Thr Lys Asp Ala
                          325 330 335
          Val Leu Ile Ala Asp Arg Asn Ala Val Gln Asp Val Lys Lys Val Val
                      340 345 350
          Glu Gln Ile Lys Ala Asp Gly Arg His Glu His Arg Val His Arg Glu
                  355 360 365
          Val Tyr Arg Pro Trp Gly Lys Tyr Asp Ser Ile Asp Ala Gly Asp Arg
              370 375 380
          Tyr Gln Val Lys Arg Ile Thr Val Lys Pro Gly Glu Gly Leu Ser Val
          385 390 395 400
          Gln Met His His His Arg Ala Glu His Trp Val Val Val Ala Gly Thr
                          405 410 415
          Ala Lys Val Thr Ile Asp Gly Asp Ile Lys Leu Leu Gly Glu Asn Glu
                      420 425 430
          Ser Ile Tyr Ile Pro Leu Gly Ala Thr His Cys Leu Glu Asn Pro Gly
                  435 440 445
          Lys Ile Pro Leu Asp Leu Ile Glu Val Arg Ser Gly Ser Tyr Leu Glu
              450 455 460
          Glu Asp Asp Val Val Arg Phe Ala Asp Arg Tyr Gly Arg Val
          465 470 475
           <![CDATA[ <210> 8]]>
           <![CDATA[ <211> 373]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Escherichia coli K-12 MG1655]]>
           <![CDATA[ <400> 8]]>
          Met Ser Lys Val Ala Leu Ile Thr Gly Val Thr Gly Gln Asp Gly Ser
          1 5 10 15
          Tyr Leu Ala Glu Phe Leu Leu Glu Lys Gly Tyr Glu Val His Gly Ile
                      20 25 30
          Lys Arg Arg Ala Ser Ser Phe Asn Thr Glu Arg Val Asp His Ile Tyr
                  35 40 45
          Gln Asp Pro His Thr Cys Asn Pro Lys Phe His Leu His Tyr Gly Asp
              50 55 60
          Leu Ser Asp Thr Ser Asn Leu Thr Arg Ile Leu Arg Glu Val Gln Pro
          65 70 75 80
          Asp Glu Val Tyr Asn Leu Gly Ala Met Ser His Val Ala Val Ser Phe
                          85 90 95
          Glu Ser Pro Glu Tyr Thr Ala Asp Val Asp Ala Met Gly Thr Leu Arg
                      100 105 110
          Leu Leu Glu Ala Ile Arg Phe Leu Gly Leu Glu Lys Lys Thr Arg Phe
                  115 120 125
          Tyr Gln Ala Ser Thr Ser Glu Leu Tyr Gly Leu Val Gln Glu Ile Pro
              130 135 140
          Gln Lys Glu Thr Thr Pro Phe Tyr Pro Arg Ser Pro Tyr Ala Val Ala
          145 150 155 160
          Lys Leu Tyr Ala Tyr Trp Ile Thr Val Asn Tyr Arg Glu Ser Tyr Gly
                          165 170 175
          Met Tyr Ala Cys Asn Gly Ile Leu Phe Asn His Glu Ser Pro Arg Arg
                      180 185 190
          Gly Glu Thr Phe Val Thr Arg Lys Ile Thr Arg Ala Ile Ala Asn Ile
                  195 200 205
          Ala Gln Gly Leu Glu Ser Cys Leu Tyr Leu Gly Asn Met Asp Ser Leu
              210 215 220
          Arg Asp Trp Gly His Ala Lys Asp Tyr Val Lys Met Gln Trp Met Met
          225 230 235 240
          Leu Gln Gln Glu Gln Pro Glu Asp Phe Val Ile Ala Thr Gly Val Gln
                          245 250 255
          Tyr Ser Val Arg Gln Phe Val Glu Met Ala Ala Ala Gln Leu Gly Ile
                      260 265 270
          Lys Leu Arg Phe Glu Gly Thr Gly Val Glu Glu Lys Gly Ile Val Val
                  275 280 285
          Ser Val Thr Gly His Asp Ala Pro Gly Val Lys Pro Gly Asp Val Ile
              290 295 300
          Ile Ala Val Asp Pro Arg Tyr Phe Arg Pro Ala Glu Val Glu Thr Leu
          305 310 315 320
          Leu Gly Asp Pro Thr Lys Ala His Glu Lys Leu Gly Trp Lys Pro Glu
                          325 330 335
          Ile Thr Leu Arg Glu Met Val Ser Glu Met Val Ala Asn Asp Leu Glu
                      340 345 350
          Ala Ala Lys Lys His Ser Leu Leu Lys Ser His Gly Tyr Asp Val Ala
                  355 360 365
          Ile Ala Leu Glu Ser
              370
           <![CDATA[ <210> 9]]>
           <![CDATA[ <211> 321]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Escherichia coli K-12 MG1655]]>
           <![CDATA[ <400> 9]]>
          Met Ser Lys Gln Arg Val Phe Ile Ala Gly His Arg Gly Met Val Gly
          1 5 10 15
          Ser Ala Ile Arg Arg Gln Leu Glu Gln Arg Gly Asp Val Glu Leu Val
                      20 25 30
          Leu Arg Thr Arg Asp Glu Leu Asn Leu Leu Asp Ser Arg Ala Val His
                  35 40 45
          Asp Phe Phe Ala Ser Glu Arg Ile Asp Gln Val Tyr Leu Ala Ala Ala
              50 55 60
          Lys Val Gly Gly Ile Val Ala Asn Asn Thr Tyr Pro Ala Asp Phe Ile
          65 70 75 80
          Tyr Gln Asn Met Met Ile Glu Ser Asn Ile Ile His Ala Ala His Gln
                          85 90 95
          Asn Asp Val Asn Lys Leu Leu Phe Leu Gly Ser Ser Cys Ile Tyr Pro
                      100 105 110
          Lys Leu Ala Lys Gln Pro Met Ala Glu Ser Glu Leu Leu Gln Gly Thr
                  115 120 125
          Leu Glu Pro Thr Asn Glu Pro Tyr Ala Ile Ala Lys Ile Ala Gly Ile
              130 135 140
          Lys Leu Cys Glu Ser Tyr Asn Arg Gln Tyr Gly Arg Asp Tyr Arg Ser
          145 150 155 160
          Val Met Pro Thr Asn Leu Tyr Gly Pro His Asp Asn Phe His Pro Ser
                          165 170 175
          Asn Ser His Val Ile Pro Ala Leu Leu Arg Arg Phe His Glu Ala Thr
                      180 185 190
          Ala Gln Asn Ala Pro Asp Val Val Val Trp Gly Ser Gly Thr Pro Met
                  195 200 205
          Arg Glu Phe Leu His Val Asp Asp Met Ala Ala Ala Ser Ile His Val
              210 215 220
          Met Glu Leu Ala His Glu Val Trp Leu Glu Asn Thr Gln Pro Met Leu
          225 230 235 240
          Ser His Ile Asn Val Gly Thr Gly Val Asp Cys Thr Ile Arg Glu Leu
                          245 250 255
          Ala Gln Thr Ile Ala Lys Val Val Gly Tyr Lys Gly Arg Val Val Phe
                      260 265 270
          Asp Ala Ser Lys Pro Asp Gly Thr Pro Arg Lys Leu Leu Asp Val Thr
                  275 280 285
          Arg Leu His Gln Leu Gly Trp Tyr His Glu Ile Ser Leu Glu Ala Gly
              290 295 300
          Leu Ala Ser Thr Tyr Gln Trp Phe Leu Glu Asn Gln Asp Arg Phe Arg
          305 310 315 320
          Gly
           <![CDATA[ <210> 10]]>
           <![CDATA[ <211> 438]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Escherichia coli K-12 MG1655]]>
           <![CDATA[ <400> 10]]>
          Met Gly Asn Thr Ser Ile Gln Thr Gln Ser Tyr Arg Ala Val Asp Lys
          1 5 10 15
          Asp Ala Gly Gln Ser Arg Ser Tyr Ile Ile Pro Phe Ala Leu Leu Cys
                      20 25 30
          Ser Leu Phe Phe Leu Trp Ala Val Ala Asn Asn Leu Asn Asp Ile Leu
                  35 40 45
          Leu Pro Gln Phe Gln Gln Ala Phe Thr Leu Thr Asn Phe Gln Ala Gly
              50 55 60
          Leu Ile Gln Ser Ala Phe Tyr Phe Gly Tyr Phe Ile Ile Pro Ile Pro
          65 70 75 80
          Ala Gly Ile Leu Met Lys Lys Leu Ser Tyr Lys Ala Gly Ile Ile Thr
                          85 90 95
          Gly Leu Phe Leu Tyr Ala Leu Gly Ala Ala Leu Phe Trp Pro Ala Ala
                      100 105 110
          Glu Ile Met Asn Tyr Thr Leu Phe Leu Val Gly Leu Phe Ile Ile Ala
                  115 120 125
          Ala Gly Leu Gly Cys Leu Glu Thr Ala Ala Asn Pro Phe Val Thr Val
              130 135 140
          Leu Gly Pro Glu Ser Ser Gly His Phe Arg Leu Asn Leu Ala Gln Thr
          145 150 155 160
          Phe Asn Ser Phe Gly Ala Ile Ile Ala Val Val Phe Gly Gln Ser Leu
                          165 170 175
          Ile Leu Ser Asn Val Pro His Gln Ser Gln Asp Val Leu Asp Lys Met
                      180 185 190
          Ser Pro Glu Gln Leu Ser Ala Tyr Lys His Ser Leu Val Leu Ser Val
                  195 200 205
          Gln Thr Pro Tyr Met Ile Ile Val Ala Ile Val Leu Leu Val Ala Leu
              210 215 220
          Leu Ile Met Leu Thr Lys Phe Pro Ala Leu Gln Ser Asp Asn His Ser
          225 230 235 240
          Asp Ala Lys Gln Gly Ser Phe Ser Ala Ser Leu Ser Arg Leu Ala Arg
                          245 250 255
          Ile Arg His Trp Arg Trp Ala Val Leu Ala Gln Phe Cys Tyr Val Gly
                      260 265 270
          Ala Gln Thr Ala Cys Trp Ser Tyr Leu Ile Arg Tyr Ala Val Glu Glu
                  275 280 285
          Ile Pro Gly Met Thr Ala Gly Phe Ala Ala Asn Tyr Leu Thr Gly Thr
              290 295 300
          Met Val Cys Phe Phe Ile Gly Arg Phe Thr Gly Thr Trp Leu Ile Ser
          305 310 315 320
          Arg Phe Ala Pro His Lys Val Leu Ala Ala Tyr Ala Leu Ile Ala Met
                          325 330 335
          Ala Leu Cys Leu Ile Ser Ala Phe Ala Gly Gly His Val Gly Leu Ile
                      340 345 350
          Ala Leu Thr Leu Cys Ser Ala Phe Met Ser Ile Gln Tyr Pro Thr Ile
                  355 360 365
          Phe Ser Leu Gly Ile Lys Asn Leu Gly Gln Asp Thr Lys Tyr Gly Ser
              370 375 380
          Ser Phe Ile Val Met Thr Ile Ile Gly Gly Gly Ile Val Thr Pro Val
          385 390 395 400
          Met Gly Phe Val Ser Asp Ala Ala Gly Asn Ile Pro Thr Ala Glu Leu
                          405 410 415
          Ile Pro Ala Leu Cys Phe Ala Val Ile Phe Ile Phe Ala Arg Phe Arg
                      420 425 430
          Ser Gln Thr Ala Thr Asn
                  435
           <![CDATA[ <210> 11]]>
           <![CDATA[ <211> 949]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Bacteroides fragilis]]>
           <![CDATA[ <400> 11]]>
          Met Gln Lys Leu Leu Ser Leu Pro Ser Asn Leu Val Gln Ser Phe His
          1 5 10 15
          Glu Leu Glu Arg Val Asn Arg Thr Asp Trp Phe Cys Thr Ser Asp Pro
                      20 25 30
          Val Gly Lys Lys Leu Gly Ser Gly Gly Gly Thr Ser Trp Leu Leu Glu
                  35 40 45
          Glu Cys Tyr Asn Glu Tyr Ser Asp Gly Ala Thr Phe Gly Glu Trp Leu
              50 55 60
          Glu Lys Glu Lys Arg Ile Leu Leu His Ala Gly Gly Gln Ser Arg Arg
          65 70 75 80
          Leu Pro Gly Tyr Ala Pro Ser Gly Lys Ile Leu Thr Pro Val Pro Val
                          85 90 95
          Phe Arg Trp Glu Arg Gly Gln His Leu Gly Gln Asn Leu Leu Ser Leu
                      100 105 110
          Gln Leu Pro Leu Tyr Glu Lys Ile Met Ser Leu Ala Pro Asp Lys Leu
                  115 120 125
          His Thr Leu Ile Ala Ser Gly Asp Val Tyr Ile Arg Ser Glu Lys Pro
              130 135 140
          Leu Gln Ser Ile Pro Glu Ala Asp Val Val Cys Tyr Gly Leu Trp Val
          145 150 155 160
          Asp Pro Ser Leu Ala Thr His His Gly Val Phe Ala Ser Asp Arg Lys
                          165 170 175
          His Pro Glu Gln Leu Asp Phe Met Leu Gln Lys Pro Ser Leu Ala Glu
                      180 185 190
          Leu Glu Ser Leu Ser Lys Thr His Leu Phe Leu Met Asp Ile Gly Ile
                  195 200 205
          Trp Leu Leu Ser Asp Arg Ala Val Glu Ile Leu Met Lys Arg Ser His
              210 215 220
          Lys Glu Ser Ser Glu Glu Leu Lys Tyr Tyr Asp Leu Tyr Ser Asp Phe
          225 230 235 240
          Gly Leu Ala Leu Gly Thr His Pro Arg Ile Glu Asp Glu Glu Val Asn
                          245 250 255
          Thr Leu Ser Val Ala Ile Leu Pro Leu Pro Gly Gly Glu Phe Tyr His
                      260 265 270
          Tyr Gly Thr Ser Lys Glu Leu Ile Ser Ser Thr Leu Ser Val Gln Asn
                  275 280 285
          Lys Val Tyr Asp Gln Arg Arg Ile Met His Arg Lys Val Lys Pro Asn
              290 295 300
          Pro Ala Met Phe Val Gln Asn Ala Val Val Arg Ile Pro Leu Cys Ala
          305 310 315 320
          Glu Asn Ala Asp Leu Trp Ile Glu Asn Ser His Ile Gly Pro Lys Trp
                          325 330 335
          Lys Ile Ala Ser Arg His Ile Ile Thr Gly Val Pro Glu Asn Asp Trp
                      340 345 350
          Ser Leu Ala Val Pro Ala Gly Val Cys Val Asp Val Val Pro Met Gly
                  355 360 365
          Asp Lys Gly Phe Val Ala Arg Pro Tyr Gly Leu Asp Asp Val Phe Lys
              370 375 380
          Gly Asp Leu Arg Asp Ser Lys Thr Thr Leu Thr Gly Ile Pro Phe Gly
          385 390 395 400
          Glu Trp Met Ser Lys Arg Gly Leu Ser Tyr Thr Asp Leu Lys Gly Arg
                          405 410 415
          Thr Asp Asp Leu Gln Ala Val Ser Val Phe Pro Met Val Asn Ser Val
                      420 425 430
          Glu Glu Leu Gly Leu Val Leu Arg Trp Met Leu Ser Glu Pro Glu Leu
                  435 440 445
          Glu Glu Gly Lys Asn Ile Trp Leu Arg Ser Glu His Phe Ser Ala Asp
              450 455 460
          Glu Ile Ser Ala Gly Ala Asn Leu Lys Arg Leu Tyr Ala Gln Arg Glu
          465 470 475 480
          Glu Phe Arg Lys Gly Asn Trp Lys Ala Leu Ala Val Asn His Glu Lys
                          485 490 495
          Ser Val Phe Tyr Gln Leu Asp Leu Ala Asp Ala Ala Glu Asp Phe Val
                      500 505 510
          Arg Leu Gly Leu Asp Met Pro Glu Leu Leu Pro Glu Asp Ala Leu Gln
                  515 520 525
          Met Ser Arg Ile His Asn Arg Met Leu Arg Ala Arg Ile Leu Lys Leu
              530 535 540
          Asp Gly Lys Asp Tyr Arg Pro Glu Glu Gln Ala Ala Phe Asp Leu Leu
          545 550 555 560
          Arg Asp Gly Leu Leu Asp Gly Ile Ser Asn Arg Lys Ser Thr Pro Lys
                          565 570 575
          Leu Asp Val Tyr Ser Asp Gln Ile Val Trp Gly Arg Ser Pro Val Arg
                      580 585 590
          Ile Asp Met Ala Gly Gly Trp Thr Asp Thr Pro Pro Tyr Ser Leu Tyr
                  595 600 605
          Ser Gly Gly Asn Val Val Asn Leu Ala Ile Glu Leu Asn Gly Gln Pro
              610 615 620
          Pro Leu Gln Val Tyr Val Lys Pro Cys Lys Asp Phe His Ile Val Leu
          625 630 635 640
          Arg Ser Ile Asp Met Gly Ala Met Glu Ile Val Ser Thr Phe Asp Glu
                          645 650 655
          Leu Gln Asp Tyr Lys Lys Lys Ile Gly Ser Pro Phe Ser Ile Pro Lys Ala
                      660 665 670
          Ala Leu Ser Leu Ala Gly Phe Ala Pro Ala Phe Ser Ala Val Ser Tyr
                  675 680 685
          Ala Ser Leu Glu Glu Gln Leu Lys Asp Phe Gly Ala Gly Ile Glu Val
              690 695 700
          Thr Leu Leu Ala Ala Ile Pro Ala Gly Ser Gly Leu Gly Thr Ser Ser
          705 710 715 720
          Ile Leu Ala Ser Thr Val Leu Gly Ala Ile Asn Asp Phe Cys Gly Leu
                          725 730 735
          Ala Trp Asp Lys Asn Glu Ile Cys Gln Arg Thr Leu Val Leu Glu Gln
                      740 745 750
          Leu Leu Thr Thr Gly Gly Gly Trp Gln Asp Gln Tyr Gly Gly Val Leu
                  755 760 765
          Gln Gly Val Lys Leu Leu Gln Thr Glu Ala Gly Phe Ala Gln Ser Pro
              770 775 780
          Leu Val Arg Trp Leu Pro Asp His Leu Phe Thr His Pro Glu Tyr Lys
          785 790 795 800
          Asp Cys His Leu Leu Tyr Tyr Thr Gly Ile Thr Arg Thr Ala Lys Gly
                          805 810 815
          Ile Leu Ala Glu Ile Val Ser Ser Met Phe Leu Asn Ser Ser Leu His
                      820 825 830
          Leu Asn Leu Leu Ser Glu Met Lys Ala His Ala Leu Asp Met Asn Glu
                  835 840 845
          Ala Ile Gln Arg Gly Ser Phe Val Glu Phe Gly Arg Leu Val Gly Lys
              850 855 860
          Thr Trp Glu Gln Asn Lys Ala Leu Asp Ser Gly Thr Asn Pro Pro Ala
          865 870 875 880
          Val Glu Ala Ile Ile Asp Leu Ile Lys Asp Tyr Thr Leu Gly Tyr Lys
                          885 890 895
          Leu Pro Gly Ala Gly Gly Gly Gly Tyr Leu Tyr Met Val Ala Lys Asp
                      900 905 910
          Pro Gln Ala Ala Val Arg Ile Arg Lys Ile Leu Thr Glu Asn Ala Pro
                  915 920 925
          Asn Pro Arg Ala Arg Phe Val Glu Met Thr Leu Ser Asp Lys Gly Phe
              930 935 940
          Gln Val Ser Arg Ser
          945
           <![CDATA[ <210> 12]]>
           <![CDATA[ <211> 417]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Escherichia coli K-12 MG1655]]>
           <![CDATA[ <400> 12]]>
          Met Tyr Tyr Leu Lys Asn Thr Asn Phe Trp Met Phe Gly Leu Phe Phe
          1 5 10 15
          Phe Phe Tyr Phe Phe Ile Met Gly Ala Tyr Phe Pro Phe Phe Pro Ile
                      20 25 30
          Trp Leu His Asp Ile Asn His Ile Ser Lys Ser Asp Thr Gly Ile Ile
                  35 40 45
          Phe Ala Ala Ile Ser Leu Phe Ser Leu Leu Phe Gln Pro Leu Phe Gly
              50 55 60
          Leu Leu Ser Asp Lys Leu Gly Leu Arg Lys Tyr Leu Leu Trp Ile Ile
          65 70 75 80
          Thr Gly Met Leu Val Met Phe Ala Pro Phe Phe Ile Phe Ile Phe Gly
                          85 90 95
          Pro Leu Leu Gln Tyr Asn Ile Leu Val Gly Ser Ile Val Gly Gly Ile
                      100 105 110
          Tyr Leu Gly Phe Cys Phe Asn Ala Gly Ala Pro Ala Val Glu Ala Phe
                  115 120 125
          Ile Glu Lys Val Ser Arg Arg Ser Asn Phe Glu Phe Gly Arg Ala Arg
              130 135 140
          Met Phe Gly Cys Val Gly Trp Ala Leu Cys Ala Ser Ile Val Gly Ile
          145 150 155 160
          Met Phe Thr Ile Asn Asn Gln Phe Val Phe Trp Leu Gly Ser Gly Cys
                          165 170 175
          Ala Leu Ile Leu Ala Val Leu Leu Phe Phe Ala Lys Thr Asp Ala Pro
                      180 185 190
          Ser Ser Ala Thr Val Ala Asn Ala Val Gly Ala Asn His Ser Ala Phe
                  195 200 205
          Ser Leu Lys Leu Ala Leu Glu Leu Phe Arg Gln Pro Lys Leu Trp Phe
              210 215 220
          Leu Ser Leu Tyr Val Ile Gly Val Ser Cys Thr Tyr Asp Val Phe Asp
          225 230 235 240
          Gln Gln Phe Ala Asn Phe Phe Thr Ser Phe Phe Ala Thr Gly Glu Gln
                          245 250 255
          Gly Thr Arg Val Phe Gly Tyr Val Thr Thr Met Gly Glu Leu Leu Asn
                      260 265 270
          Ala Ser Ile Met Phe Phe Ala Pro Leu Ile Ile Asn Arg Ile Gly Gly
                  275 280 285
          Lys Asn Ala Leu Leu Leu Ala Gly Thr Ile Met Ser Val Arg Ile Ile
              290 295 300
          Gly Ser Ser Phe Ala Thr Ser Ala Leu Glu Val Val Ile Leu Lys Thr
          305 310 315 320
          Leu His Met Phe Glu Val Pro Phe Leu Leu Val Gly Cys Phe Lys Tyr
                          325 330 335
          Ile Thr Ser Gln Phe Glu Val Arg Phe Ser Ala Thr Ile Tyr Leu Val
                      340 345 350
          Cys Phe Cys Phe Phe Lys Gln Leu Ala Met Ile Phe Met Ser Val Leu
                  355 360 365
          Ala Gly Asn Met Tyr Glu Ser Ile Gly Phe Gln Gly Ala Tyr Leu Val
              370 375 380
          Leu Gly Leu Val Ala Leu Gly Phe Thr Leu Ile Ser Val Phe Thr Leu
          385 390 395 400
          Ser Gly Pro Gly Pro Leu Ser Leu Leu Arg Arg Gln Val Asn Glu Val
                          405 410 415
          Ala
           <![CDATA[ <210> 13]]>
           <![CDATA[ <211> 275]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Neisseria meningitidis MC58]]>
           <![CDATA[ <400> 13]]>
          Met Gln Asn His Val Ile Ser Leu Ala Ser Ala Ala Glu Arg Arg Ala
          1 5 10 15
          His Ile Ala Asp Thr Phe Gly Arg His Gly Ile Pro Phe Gln Phe Phe
                      20 25 30
          Asp Ala Leu Met Pro Ser Glu Arg Leu Glu Gln Ala Met Ala Glu Leu
                  35 40 45
          Val Pro Gly Leu Ser Ala His Pro Tyr Leu Ser Gly Val Glu Lys Ala
              50 55 60
          Cys Phe Met Ser His Ala Val Leu Trp Lys Gln Ala Leu Asp Glu Gly
          65 70 75 80
          Leu Pro Tyr Ile Thr Val Phe Glu Asp Asp Val Leu Leu Gly Glu Gly
                          85 90 95
          Ala Glu Lys Phe Leu Ala Glu Asp Ala Trp Leu Gln Glu Arg Phe Asp
                      100 105 110
          Pro Asp Thr Ala Phe Ile Val Arg Leu Glu Thr Met Phe Met His Val
                  115 120 125
          Leu Thr Ser Pro Ser Gly Val Ala Asp Tyr Cys Gly Arg Ala Phe Pro
              130 135 140
          Leu Leu Glu Ser Glu His Trp Gly Thr Ala Gly Tyr Ile Ile Ser Arg
          145 150 155 160
          Lys Ala Met Arg Phe Phe Leu Asp Arg Phe Ala Ala Leu Pro Pro Glu
                          165 170 175
          Gly Leu His Pro Val Asp Leu Met Met Phe Ser Asp Phe Phe Asp Arg
                      180 185 190
          Glu Gly Met Pro Val Cys Gln Leu Asn Pro Ala Leu Cys Ala Gln Glu
                  195 200 205
          Leu His Tyr Ala Lys Phe His Asp Gln Asn Ser Ala Leu Gly Ser Leu
              210 215 220
          Ile Glu His Asp Arg Leu Leu Asn Arg Lys Gln Gln Arg Arg Asp Ser
          225 230 235 240
          Pro Ala Asn Thr Phe Lys His Arg Leu Ile Arg Ala Leu Thr Lys Ile
                          245 250 255
          Ser Arg Glu Arg Glu Lys Arg Arg Gln Arg Arg Glu Gln Phe Ile Val
                      260 265 270
          Pro Phe Gln
                  275
           <![CDATA[ <210> 14]]>
           <![CDATA[ <211> 338]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Escherichia coli K-12 MG1655]]>
           <![CDATA[ <400> 14]]>
          Met Arg Val Leu Val Thr Gly Gly Ser Gly Tyr Ile Gly Ser His Thr
          1 5 10 15
          Cys Val Gln Leu Leu Gln Asn Gly His Asp Val Ile Ile Leu Asp Asn
                      20 25 30
          Leu Cys Asn Ser Lys Arg Ser Val Leu Pro Val Ile Glu Arg Leu Gly
                  35 40 45
          Gly Lys His Pro Thr Phe Val Glu Gly Asp Ile Arg Asn Glu Ala Leu
              50 55 60
          Met Thr Glu Ile Leu His Asp His Ala Ile Asp Thr Val Ile His Phe
          65 70 75 80
          Ala Gly Leu Lys Ala Val Gly Glu Ser Val Gln Lys Pro Leu Glu Tyr
                          85 90 95
          Tyr Asp Asn Asn Val Asn Gly Thr Leu Arg Leu Ile Ser Ala Met Arg
                      100 105 110
          Ala Ala Asn Val Lys Asn Phe Ile Phe Ser Ser Ser Ala Thr Val Tyr
                  115 120 125
          Gly Asp Gln Pro Lys Ile Pro Tyr Val Glu Ser Phe Pro Thr Gly Thr
              130 135 140
          Pro Gln Ser Pro Tyr Gly Lys Ser Lys Leu Met Val Glu Gln Ile Leu
          145 150 155 160
          Thr Asp Leu Gln Lys Ala Gln Pro Asp Trp Ser Ile Ala Leu Leu Arg
                          165 170 175
          Tyr Phe Asn Pro Val Gly Ala His Pro Ser Gly Asp Met Gly Glu Asp
                      180 185 190
          Pro Gln Gly Ile Pro Asn Asn Leu Met Pro Tyr Ile Ala Gln Val Ala
                  195 200 205
          Val Gly Arg Arg Asp Ser Leu Ala Ile Phe Gly Asn Asp Tyr Pro Thr
              210 215 220
          Glu Asp Gly Thr Gly Val Arg Asp Tyr Ile His Val Met Asp Leu Ala
          225 230 235 240
          Asp Gly His Val Val Ala Met Glu Lys Leu Ala Asn Lys Pro Gly Val
                          245 250 255
          His Ile Tyr Asn Leu Gly Ala Gly Val Gly Asn Ser Val Leu Asp Val
                      260 265 270
          Val Asn Ala Phe Ser Lys Ala Cys Gly Lys Pro Val Asn Tyr His Phe
                  275 280 285
          Ala Pro Arg Arg Glu Gly Asp Leu Pro Ala Tyr Trp Ala Asp Ala Ser
              290 295 300
          Lys Ala Asp Arg Glu Leu Asn Trp Arg Val Thr Arg Thr Leu Asp Glu
          305 310 315 320
          Met Ala Gln Asp Thr Trp His Trp Gln Ser Arg His Pro Gln Gly Tyr
                          325 330 335
          Pro Asp
           <![CDATA[ <210> 15]]>
           <![CDATA[ <211> 587]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Kluyveromyces lactis]]>
           <![CDATA[ <400> 15]]>
          Met Ala Asp His Ser Ser Ser Ser Ser Ser Ser Leu Gln Lys Lys Pro Ile
          1 5 10 15
          Asn Thr Ile Glu His Lys Asp Thr Leu Gly Asn Asp Arg Asp His Lys
                      20 25 30
          Glu Ala Leu Asn Ser Asp Asn Asp Asn Thr Ser Gly Leu Lys Ile Asn
                  35 40 45
          Gly Val Pro Ile Glu Asp Ala Arg Glu Glu Val Leu Leu Pro Gly Tyr
              50 55 60
          Leu Ser Lys Gln Tyr Tyr Lys Leu Tyr Gly Leu Cys Phe Ile Thr Tyr
          65 70 75 80
          Leu Cys Ala Thr Met Gln Gly Tyr Asp Gly Ala Leu Met Gly Ser Ile
                          85 90 95
          Tyr Thr Glu Asp Ala Tyr Leu Lys Tyr Tyr His Leu Asp Ile Asn Ser
                      100 105 110
          Ser Ser Gly Thr Gly Leu Val Phe Ser Ile Phe Asn Val Gly Gln Ile
                  115 120 125
          Cys Gly Ala Phe Phe Val Pro Leu Met Asp Trp Lys Gly Arg Lys Pro
              130 135 140
          Ala Ile Leu Ile Gly Cys Leu Gly Val Val Ile Gly Ala Ile Ile Ser
          145 150 155 160
          Ser Leu Thr Thr Thr Lys Ser Ala Leu Ile Gly Gly Arg Trp Phe Val
                          165 170 175
          Ala Phe Phe Ala Thr Ile Ala Asn Ala Ala Ala Pro Thr Tyr Cys Ala
                      180 185 190
          Glu Val Ala Pro Ala His Leu Arg Gly Lys Val Ala Gly Leu Tyr Asn
                  195 200 205
          Thr Leu Trp Ser Val Gly Ser Ile Val Ala Ala Phe Ser Thr Tyr Gly
              210 215 220
          Thr Asn Lys Asn Phe Pro Asn Ser Ser Lys Ala Phe Lys Ile Pro Leu
          225 230 235 240
          Tyr Leu Gln Met Met Phe Pro Gly Leu Val Cys Ile Phe Gly Trp Leu
                          245 250 255
          Ile Pro Glu Ser Pro Arg Trp Leu Val Gly Val Gly Arg Glu Glu Glu
                      260 265 270
          Ala Arg Glu Phe Ile Ile Lys Tyr His Leu Asn Gly Asp Arg Thr His
                  275 280 285
          Pro Leu Leu Asp Met Glu Met Ala Glu Ile Ile Glu Ser Phe His Gly
              290 295 300
          Thr Asp Leu Ser Asn Pro Leu Glu Met Leu Asp Val Arg Ser Leu Phe
          305 310 315 320
          Arg Thr Arg Ser Asp Arg Tyr Arg Ala Met Leu Val Ile Leu Met Ala
                          325 330 335
          Trp Phe Gly Gln Phe Ser Gly Asn Asn Val Cys Ser Tyr Tyr Leu Pro
                      340 345 350
          Thr Met Leu Arg Asn Val Gly Met Lys Ser Val Ser Leu Asn Val Leu
                  355 360 365
          Met Asn Gly Val Tyr Ser Ile Val Thr Trp Ile Ser Ser Ile Cys Gly
              370 375 380
          Ala Phe Phe Ile Asp Lys Ile Gly Arg Arg Glu Gly Phe Leu Gly Ser
          385 390 395 400
          Ile Ser Gly Ala Ala Leu Ala Leu Thr Gly Leu Ser Ile Cys Thr Ala
                          405 410 415
          Arg Tyr Glu Lys Thr Lys Lys Lys Lys Ser Ala Ser Asn Gly Ala Leu Val
                      420 425 430
          Phe Ile Tyr Leu Phe Gly Gly Ile Phe Ser Phe Ala Phe Thr Pro Met
                  435 440 445
          Gln Ser Met Tyr Ser Thr Glu Val Ser Thr Asn Leu Thr Arg Ser Lys
              450 455 460
          Ala Gln Leu Leu Asn Phe Val Val Ser Gly Val Ala Gln Phe Val Asn
          465 470 475 480
          Gln Phe Ala Thr Pro Lys Ala Met Lys Asn Ile Lys Tyr Trp Phe Tyr
                          485 490 495
          Val Phe Tyr Val Phe Phe Asp Ile Phe Glu Phe Ile Val Ile Tyr Phe
                      500 505 510
          Phe Phe Val Glu Thr Lys Gly Arg Ser Leu Glu Glu Leu Glu Val Val
                  515 520 525
          Phe Glu Ala Pro Asn Pro Arg Lys Ala Ser Val Asp Gln Ala Phe Leu
              530 535 540
          Ala Gln Val Arg Ala Thr Leu Val Gln Arg Asn Asp Val Arg Val Ala
          545 550 555 560
          Asn Ala Gln Asn Leu Lys Glu Gln Glu Pro Leu Lys Ser Asp Ala Asp
                          565 570 575
          His Val Glu Lys Leu Ser Glu Ala Glu Ser Val
                      580 585
           <![CDATA[ <210> 16]]>
           <![CDATA[ <211> 299]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Helicobacter pylori]]>
           <![CDATA[ <400> 16]]>
          Met Ala Phe Lys Val Val Gln Ile Cys Gly Gly Leu Gly Asn Gln Met
          1 5 10 15
          Phe Gln Tyr Ala Phe Ala Lys Ser Leu Gln Lys His Ser Asn Thr Pro
                      20 25 30
          Val Leu Leu Asp Ile Thr Ser Phe Asp Trp Ser Asn Arg Lys Met Gln
                  35 40 45
          Leu Glu Leu Phe Pro Ile Asp Leu Pro Tyr Ala Ser Glu Lys Glu Ile
              50 55 60
          Ala Ile Ala Lys Met Gln His Leu Pro Lys Leu Val Arg Asn Val Leu
          65 70 75 80
          Lys Cys Met Gly Phe Asp Arg Val Ser Gln Glu Ile Val Phe Glu Tyr
                          85 90 95
          Glu Pro Lys Leu Leu Lys Thr Ser Arg Leu Thr Tyr Phe Tyr Gly Tyr
                      100 105 110
          Phe Gln Asp Pro Arg Tyr Phe Asp Ala Ile Ser Pro Leu Ile Lys Gln
                  115 120 125
          Thr Phe Thr Leu Pro Pro Pro Pro Glu Asn Gly Asn Asn Lys Lys Lys
              130 135 140
          Glu Glu Glu Tyr His Arg Lys Leu Ala Leu Ile Leu Ala Ala Lys Asn
          145 150 155 160
          Ser Val Phe Val His Ile Arg Arg Gly Asp Tyr Val Gly Ile Gly Cys
                          165 170 175
          Gln Leu Gly Ile Asp Tyr Gln Lys Lys Ala Leu Glu Tyr Met Ala Lys
                      180 185 190
          Arg Val Pro Asn Met Glu Leu Phe Val Phe Cys Glu Asp Leu Glu Phe
                  195 200 205
          Thr Gln Asn Leu Asp Leu Gly Tyr Pro Phe Met Asp Met Thr Thr Arg
              210 215 220
          Asp Lys Glu Glu Glu Ala Tyr Trp Asp Met Leu Leu Met Gln Ser Cys
          225 230 235 240
          Lys His Gly Ile Ile Ala Asn Ser Thr Tyr Ser Trp Trp Ala Ala Tyr
                          245 250 255
          Leu Ile Asn Asn Pro Glu Lys Ile Ile Ile Gly Pro Lys His Trp Leu
                      260 265 270
          Phe Gly His Glu Asn Ile Leu Cys Lys Glu Trp Val Lys Ile Glu Ser
                  275 280 285
          His Phe Glu Val Lys Ser Gln Lys Tyr Asn Ala
              290 295
           <![CDATA[ <210> 17]]>
           <![CDATA[ <211> 311]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Helicobacter sp. MIT 01-6242]]>
           <![CDATA[ <400> 17]]>
          Met Phe Ser Val Arg Leu Met Gly Gly Leu Gly Asn Gln Met Phe Ile
          1 5 10 15
          Tyr Ala Phe Ala Lys Ala Ile Lys Ala Gln Gly Tyr Pro Val Arg Leu
                      20 25 30
          Phe Tyr Tyr Asp Thr Asp Tyr Asn Val Pro Gln Thr His Asn Ile Arg
                  35 40 45
          Asn Leu Glu Ile Val Asp Phe Gly Ile Ala Met Cys Ile Glu Thr Met
              50 55 60
          Cys Tyr Glu Glu Pro Gln Ile Lys Lys Ser Phe Phe Glu Arg Ala Leu
          65 70 75 80
          Gly Phe Ile Lys Arg Lys Leu Lys Ile His Ser Pro His Ser Ser Ser
                          85 90 95
          Leu Ile Ser Asp His Cys Glu Ile Ala Leu Thr Lys Asp Phe Leu Asp
                      100 105 110
          Thr Leu Asn Pro Asn Ala Met Phe Asn Gly Tyr Phe Gln Asn Val Val
                  115 120 125
          Phe Phe Asp His Leu Arg Glu Ser Leu Leu Arg Asp Phe Thr Leu Lys
              130 135 140
          Arg Pro Leu Thr Pro Ala Asn Glu Ala Leu Lys His Gln Ile Leu Gln
          145 150 155 160
          Thr Pro Asn Ser Cys Phe Leu His Ile Arg Arg Gly Asp Tyr Leu Gln
                          165 170 175
          Ile Pro Ile Tyr Val Lys Leu Gly Ser Thr Tyr Tyr Asn Asn Ala Ile
                      180 185 190
          Lys Ala Leu Lys Asp Lys Ile Ser Lys Pro His Ile Phe Val Phe Ser
                  195 200 205
          Asn Asp Ile Ala Trp Cys Lys Glu Phe Phe Leu Asp Ser Leu Asp Pro
              210 215 220
          Leu Val Ile Glu Asn Val Thr Phe Ser Phe Ile Glu Asn Asn Asp Glu
          225 230 235 240
          Gly Asn Ala Ile Glu Glu Met Glu Leu Met Arg Ser Cys Gln His Ala
                          245 250 255
          Ile Ile Ala Asn Ser Thr Phe Ser Trp Trp Ala Ala Tyr Leu Ile Asp
                      260 265 270
          Ser Ala Gln Lys Leu Cys Ile Met Pro Lys His Phe Phe Asn Asp Pro
                  275 280 285
          Gln Gln Glu Val Ala His Lys Leu Ile Pro Pro Pro Leu His Ser Leu
              290 295 300
          Ser Gln Thr Ile Val Ile Gly
          305 310
           <![CDATA[ <210> 18]]>
           <![CDATA[ <211> 261]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Akkermansia muciniphila]]>
           <![CDATA[ <400> 18]]>
          Met Asn Met Glu Arg Lys Thr Gly Leu Met Asn Lys Lys Tyr Val Ser
          1 5 10 15
          Pro Cys Phe Leu Pro Gly Met Arg Leu Gly Asn Ile Met Phe Thr Leu
                      20 25 30
          Ala Ala Ala Cys Ala His Ala Arg Thr Val Gly Val Glu Cys Arg Val
                  35 40 45
          Pro Trp Ala Tyr Asn Asp Ala Ser Leu Met Leu Arg Ser Arg Leu Gly
              50 55 60
          Gly Trp Val Leu Pro Ser Thr Pro Cys Gly Thr Asn Glu Pro Ser
          65 70 75 80
          Trp Gln Glu Pro Ser Phe Ala Tyr Cys Pro Val Pro Ser Arg Ile Arg
                          85 90 95
          Thr Gly Gly Leu Arg Gly Tyr Phe Gln Ser Ala Arg Tyr Phe Glu Gly
                      100 105 110
          Gln Glu Ala Phe Ile Arg Ala Leu Phe Ala Pro Leu Thr Ala Glu Lys
                  115 120 125
          Glu Pro Gly Ala Val Gly Ile His Ile Arg Leu Gly Asp Tyr Arg Arg
              130 135 140
          Leu Arg Asp Lys His Arg Ile Leu Asp Pro Gly Phe Leu Arg Arg Ala
          145 150 155 160
          Ala Gly His Leu Ser Ser Gly Lys Asn Arg Leu Val Leu Phe Ser Asp
                          165 170 175
          Glu Pro Asp Glu Ala Ala Glu Met Leu Ala Arg Val Pro Ala Phe Gly
                      180 185 190
          Arg Phe Ala Leu Glu Ile Asp Arg Gly Ala Pro Cys Glu Ser Leu Arg
                  195 200 205
          Arg Met Thr Ala Met Glu Glu Leu Val Met Ser Cys Ser Ser Phe Ser
              210 215 220
          Trp Trp Gly Ala Trp Leu Gly Asn Thr Arg Lys Val Ile Val Pro Arg
          225 230 235 240
          Asp Trp Phe Val Gly Gly Val Glu Asp Tyr Arg Asp Ile Tyr Leu Pro
                          245 250 255
          His Trp Val Thr Leu
                      260
           <![CDATA[ <210> 19]]>
           <![CDATA[ <211> 297]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Pyrromonas catarrhalis]]>
           <![CDATA[ <400> 19]]>
          Met Lys Arg Ile Tyr Leu Ser Ile Tyr Gly Gly Leu Gly Asn Gln Leu
          1 5 10 15
          Tyr Ile Leu Ala Tyr Ala Asp Tyr Leu Gln Arg Met Leu Gly Thr Arg
                      20 25 30
          Pro Tyr Leu Leu Asn Glu Leu Gln Arg Thr Lys Ala Asp Thr Ser Ser
                  35 40 45
          Leu Asp Arg Thr Arg Arg Asp Leu Phe Ser Glu Leu Ile Ala Tyr Leu
              50 55 60
          Gly Phe Gln Phe Val Asp Thr Asp Ser Arg Glu Phe Arg Leu Leu Lys
          65 70 75 80
          Lys Trp Glu Arg His Ile Lys His Tyr Gln Glu Glu Pro Asn Lys His
                          85 90 95
          Gly Ile Tyr Leu Gln Asn Ile Leu Pro Pro Leu Gln Glu Asn His Arg
                      100 105 110
          Trp Thr Leu Pro Leu Val Cys Arg Val Ser Gly Tyr Phe Gln Ser Cys
                  115 120 125
          His Tyr Val Gly Thr Asp Phe Arg Met Arg Val Ser Lys Phe Leu Glu
              130 135 140
          Arg His Ala Thr Ser Ala Asp Leu Val Arg Glu Tyr Thr Ser Met Ile
          145 150 155 160
          Gln Pro Glu Asp Val Ala Ile His Leu Arg Arg Gly Asp Phe Val Ala
                          165 170 175
          Leu Gln His Thr Gly Ile Gln Leu Phe Gly Ala Glu His Tyr Thr Lys
                      180 185 190
          Gly Leu Ala Leu Gln Ala Gln Gln Gln Pro Ile Gly Arg Val Phe Val
                  195 200 205
          Phe Ser Asp Asp Phe Glu Ala Ile Gly Glu Glu Leu Ser Leu Leu Ala
              210 215 220
          Asn Asn Tyr Gln Leu Val Leu Val Lys Gly Leu Thr Pro Leu Gln Asp
          225 230 235 240
          Leu Phe Leu Leu Thr Cys Phe Arg Arg Tyr Val Leu Ala Asn Ser Thr
                          245 250 255
          Phe Ser Trp Trp Gly Ala Leu Cys Ser Lys Tyr Gly Asp Ala Val Lys
                      260 265 270
          Val Val Val Pro Lys Lys Pro Leu Leu Ile Ser Tyr Pro Glu Asp Ser
                  275 280 285
          Tyr Phe Pro Pro Ser Trp Glu Gln Ile
              290 295
           <![CDATA[ <210> 20]]>
           <![CDATA[ <211> 289]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Capnocytophaga canis]]>
           <![CDATA[ <400> 20]]>
          Met Met Arg His Tyr Ile Cys Leu Gln Gly Gly Leu Gly Asn Gln Leu
          1 5 10 15
          Tyr Ile Leu Gly Tyr Ala Phe Tyr Leu Gln Glu Gln Gly Cys Lys Asn
                      20 25 30
          Ile Cys Leu Phe Tyr Glu Gln Lys Gln Asn Gly Asp Thr Val Asp Thr
                  35 40 45
          Gln Lys Arg Asn Ile Ile Asp Asp Leu Pro Lys Glu Leu Gly Phe Lys
              50 55 60
          Ser Leu Tyr Ile Ser Ser Leu Trp Leu Lys Val Leu Arg Asn Leu Cys
          65 70 75 80
          Lys Leu Pro Met Ile Ser Lys Leu Val Asp Tyr Tyr Glu Glu Pro Thr
                          85 90 95
          Glu Glu Trp Ala Val Phe Thr Pro Phe Ser Pro Leu Arg Lys Lys Arg
                      100 105 110
          Val Ser Val His Ile Gly Tyr Tyr Gln Ser Phe Tyr Tyr Gln Thr Pro
                  115 120 125
          Leu Phe Ile Glu Arg Leu Lys Arg Phe Phe Phe Lys Asn Val Leu Leu
              130 135 140
          Glu Arg Phe Ser Pro Val Glu Asn Asp Ala Ala Ile His Ile Arg Arg
          145 150 155 160
          Gly Asp Phe Leu Thr Gly Val Asn Thr Met Ile Tyr Ser Glu Ile Gly
                          165 170 175
          Val Lys Tyr Tyr Leu Glu Gly Leu Glu Lys Leu Asn Arg Glu Gln Gly
                      180 185 190
          Ile Gly Lys Ile Tyr Val Phe Ser Asp Asp Phe Gln Ala Ile Thr Asn
                  195 200 205
          Asp Ile Glu Glu Ile Ser Lys Ile Tyr Thr Val Glu Leu Met Gln Gly
              210 215 220
          Asn Ser Val Leu Gln Asp Ile Arg Met Leu Met Cys Phe Lys Arg Tyr
          225 230 235 240
          Val Leu Gly Asn Ser Thr Phe Ala Trp Trp Gly Ala Lys Leu Ser Glu
                          245 250 255
          Lys Gln Asn Pro Ile Val Val Val Pro Ala Thr Pro Trp Lys Ile Asn
                      260 265 270
          Leu Lys Lys Glu Val Thr Pro Tyr Pro Lys Asp Trp Ile Leu Leu Glu
                  275 280 285
          Asn
           <![CDATA[ <210> 21]]>
           <![CDATA[ <211> 289]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Capnocytophila Lidbitter's]]>
           <![CDATA[ <400> 21]]>
          Met Lys Ala Asn Tyr Val Leu Phe Gln Gly Gly Leu Gly Asn Gln Leu
          1 5 10 15
          Tyr Gln Leu Ala Tyr Thr Asp Phe Leu Lys Arg Asn Gly Tyr Ser Asn
                      20 25 30
          Val Lys Leu Ile Thr Pro Ser Asn Lys Asn Lys Gly Asp Thr Lys Asp
                  35 40 45
          Lys Asn Lys Arg Pro Leu Ile Thr Gln Leu Pro Glu Lys Ile Gly Ile
              50 55 60
          Asp Cys Val Asp Phe Gly His Lys Tyr Phe Tyr Ser Phe Leu Gln Arg
          65 70 75 80
          Leu Pro Lys Phe Pro Leu Tyr Lys Ser Phe Leu Arg Arg Leu Ile Asn
                          85 90 95
          Val Glu Lys Glu Pro Pro Tyr Gln Trp Ala Ile Phe His Pro Ile Thr
                      100 105 110
          Arg Glu Lys Tyr Arg Leu Asn Ile His Ile Gly Tyr Tyr Gln Ser Asn
                  115 120 125
          Leu Tyr Ile Ser Asp Ser Phe Lys Gln Gln Val Ala Lys Val Ile Glu
              130 135 140
          Ser Leu Ser Pro Asn Ile Lys Phe Ser Ile Thr Asn Asn Asp Val Ala
          145 150 155 160
          Ile His Ile Arg Arg Gly Asp Phe Leu Ile Gly Asn Asn Ala Ser Val
                          165 170 175
          Phe Asn Lys Ile Glu Leu Pro His Tyr Leu Gln Gly Leu Thr Ile Leu
                      180 185 190
          Ser Glu Arg Met Asn Ile Gln Lys Val Tyr Ile Phe Ser Asp Asp Phe
                  195 200 205
          Glu Ala Ile Lys Glu Asp Ile Lys Thr Ile Ala Glu Asn Tyr Glu Val
              210 215 220
          Val Leu Val Glu Gly Gln Ser Val Leu Ala Asp Phe Ala Leu Leu Gln
          225 230 235 240
          Lys Phe Thr Asn Phe Val Ile Gly Asn Ser Thr Phe Ala Trp Trp Gly
                          245 250 255
          Ala Met Leu Ala Asn Ala Ser Asn Val Ile Val Pro Lys Lys Pro Trp
                      260 265 270
          Lys Ile Glu Met Glu Asn Met Ser Pro Tyr Pro Asp Asn Trp Thr Thr
                  275 280 285
          Ile
           <![CDATA[ <210> 22]]>
           <![CDATA[ <211> 303]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Helicobacter sp. MIT 01-6242]]>
           <![CDATA[ <400> 22]]>
          Met Phe Ser Val Arg Leu Met Gly Gly Leu Gly Asn Gln Met Phe Ile
          1 5 10 15
          Tyr Ala Phe Ala Lys Ala Ile Lys Ala Gln Gly Tyr Pro Val Arg Leu
                      20 25 30
          Phe Tyr Tyr Asp Thr Asp Tyr Asn Val Pro Gln Thr His Asn Ile Arg
                  35 40 45
          Asn Leu Glu Ile Val Asp Phe Gly Leu Asn Leu Pro Ile Gln Arg Leu
              50 55 60
          Cys Ser Lys Gly Ile His Arg Asn Val Lys Arg Ile Ile Ala Phe Ile
          65 70 75 80
          His Arg Lys Ile Ala Lys Tyr Leu Phe Thr Phe Val Ser Asp Cys His
                          85 90 95
          Asn Val Ser Pro Thr Lys Asp Phe Leu Asp Thr Leu Asn Pro Asn Ala
                      100 105 110
          Met Phe Asn Gly Tyr Phe Gln Asn Val Val Phe Phe Asp His Leu Arg
                  115 120 125
          Glu Ser Leu Leu Arg Asp Phe Thr Leu Lys Arg Pro Leu Thr Pro Ala
              130 135 140
          Asn Glu Ala Leu Lys His Gln Ile Leu Gln Thr Pro Asn Ser Cys Phe
          145 150 155 160
          Leu His Ile Arg Arg Gly Asp Phe Leu Pro Leu Pro Glu Tyr Ile Gln
                          165 170 175
          Leu Asp Ser Thr Ala Tyr Tyr Asn Asn Ala Ile Lys Ala Leu Lys Asp
                      180 185 190
          Lys Ile Ser Lys Pro His Ile Phe Val Phe Ser Asn Asp Ile Ala Trp
                  195 200 205
          Cys Lys Glu Phe Phe Leu Asp Ser Leu Asp Pro Leu Val Ile Glu Asn
              210 215 220
          Val Thr Phe Ser Phe Val Glu Asn Asn Asp Glu Gly Asn Ala Ile Glu
          225 230 235 240
          Glu Met Glu Leu Met Arg Ser Cys Gln His Ala Ile Ile Ala Asn Ser
                          245 250 255
          Thr Phe Ser Trp Trp Ala Ala Tyr Leu Ile Asp Ser Ala Gln Lys Leu
                      260 265 270
          Cys Ile Met Pro Gln Arg Phe Tyr Gly Ile Gln Gln Glu Glu Asn Cys
                  275 280 285
          Asp Ile Pro Pro Pro Pro His Thr Leu Ala Val Tyr Ser Lys Asn
              290 295 300
           <![CDATA[ <210> 23]]>
           <![CDATA[ <211> 288]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Goldella denitrification]]>
           <![CDATA[ <400> 23]]>
          Met Ala Lys His Tyr Ile His Phe Ala Asn Gly Leu Gly Leu Gly Asn
          1 5 10 15
          Gln Leu Tyr Leu Leu Ala Tyr Thr Asp Tyr Leu Lys Ser Leu Gly Ala
                      20 25 30
          Asp Ala Ser Leu Phe Leu Met Gly Asn Gly Gln Val Gly Asp Thr Lys
                  35 40 45
          Asp Lys Ala Lys Arg Asn Leu Val Leu Glu Ile Pro Gln Arg Leu Gly
              50 55 60
          Met Ala Val Val Asp Lys Lys Gln Ile Ser Arg Ser Phe Arg Tyr Ile
          65 70 75 80
          Phe Cys Lys Ala Phe Tyr Gly Lys Glu Lys Leu Asn Trp Arg Glu Pro
                          85 90 95
          Lys Asp Gln Trp Ala Val Phe Phe Glu Gly Val His Gly Lys Tyr Pro
                      100 105 110
          Ile Asn Phe Tyr Tyr Gly Tyr Phe Gln Ser His His Tyr Ile Ala Glu
                  115 120 125
          Pro Phe Lys Gln Arg Leu Lys Thr Val Leu Arg Ala Leu Lys Pro Val
              130 135 140
          Asp Val Ala Val Gly Glu Asn Asp Val Ala Leu His Ile Arg Arg Gly
          145 150 155 160
          Asp Phe Leu Asn Pro Glu Asn Ala Gly Leu Ile Arg Thr Val Gly Leu
                          165 170 175
          Asp Tyr Tyr Leu Asn Gly Leu Asp Tyr Ile Arg Gln Arg Gln Thr Ile
                      180 185 190
          Gly Thr Val Tyr Ile Phe Ser Asp Asp Phe Ala Ala Ile Glu Lys Glu
                  195 200 205
          Ile Ala Ala Ile Ser Ala Asp Tyr Pro Val Arg Leu Met Gln Gly Gln
              210 215 220
          Ser Val Leu Glu Asp Met Asn Trp Leu Thr Tyr Phe Lys His Tyr Val
          225 230 235 240
          Val Gly Asn Ser Thr Phe Ala Trp Trp Gly Cys Leu Leu Ser Asp His
                          245 250 255
          Gln Asp Gly Leu Val Val Val Pro Gln Arg Pro Trp Val Gln Glu Gln
                      260 265 270
          Pro Ala Ala Ser Gln Tyr Leu Pro His Trp Val Gln Leu Pro Asn Glu
                  275 280 285
           <![CDATA[ <210> 24]]>
           <![CDATA[ <211> 288]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Pseudoalteromonas unique]]>
           <![CDATA[ <400> 24]]>
          Met Ile Lys Val Lys Ala Ile Gly Gly Leu Gly Asn Gln Leu Phe Gln
          1 5 10 15
          Tyr Ala Thr Ala Arg Ala Ile Ala Glu Lys Arg Gly Asp Gly Val Val
                      20 25 30
          Val Asp Met Ser Asp Phe Ser Ser Tyr Lys Thr His Pro Phe Cys Leu
                  35 40 45
          Asn Lys Phe Arg Cys Lys Lys Ala Thr Tyr Glu Ser Lys Pro Lys Leu Ile
              50 55 60
          Asn Lys Leu Leu Ser Asn Glu Lys Ile Arg Asn Leu Leu Gln Lys Leu
          65 70 75 80
          Gly Phe Ile Lys Lys Tyr Tyr Phe Glu Thr Gln Leu Pro Phe Asn Glu
                          85 90 95
          Asp Val Leu Leu Asn Asn Ser Ile Asn Tyr Leu Thr Gly Tyr Phe Gln
                      100 105 110
          Ser Glu Lys Tyr Phe Leu Ser Ile Arg Glu Cys Leu Leu Asp Glu Leu
                  115 120 125
          Thr Leu Ile Glu Asp Leu Asn Ile Ala Glu Thr Ala Val Ser Lys Ala
              130 135 140
          Ile Lys Asn Ala Lys Asn Ser Ile Ser Ile His Ile Arg Arg Gly Asp
          145 150 155 160
          Tyr Val Ser Asn Glu Gly Ala Asn Lys Thr His Gly Val Cys Asp Ser
                          165 170 175
          Asp Tyr Phe Lys Lys Ala Leu Asn Tyr Phe Ser Glu Arg Lys Leu Leu
                      180 185 190
          Asp Glu His Thr Glu Leu Phe Ile Phe Ser Asp Asp Ile Glu Trp Cys
                  195 200 205
          Arg Asn Asn Leu Ser Phe Asp Tyr Lys Met Asn Phe Val Asp Gly Ser
              210 215 220
          Ser Glu Arg Pro Glu Val Asp Met Val Leu Met Ser Gln Cys Lys His
          225 230 235 240
          Gln Val Ile Ser Asn Ser Thr Phe Ser Trp Trp Gly Ala Trp Leu Asn
                          245 250 255
          Lys Asn Asp Glu Lys Val Val Val Ala Pro Lys Glu Trp Phe Lys Ser
                      260 265 270
          Thr Asp Leu Asp Ser Thr Asp Ile Val Pro Asn Gln Trp Ile Lys Leu
                  275 280 285
           <![CDATA[ <210> 25]]>
           <![CDATA[ <211> 297]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Pyrromonas catarrhalis F0037]]>
           <![CDATA[ <400> 25]]>
          Met Lys Arg Ile Tyr Leu Ser Ile Tyr Gly Gly Leu Gly Asn Gln Leu
          1 5 10 15
          Tyr Ile Leu Ala Tyr Ala Asp Tyr Leu Gln Arg Met Leu Gly Thr Arg
                      20 25 30
          Pro Tyr Leu Leu Asn Glu Leu Gln Arg Thr Lys Ala Asp Thr Ser Ser
                  35 40 45
          Leu Asp Arg Thr Arg Arg Asp Leu Phe Ser Glu Leu Ile Ala Tyr Leu
              50 55 60
          Gly Phe Gln Phe Ile Asp Thr Asp Ser Arg Glu Phe Arg Leu Leu Lys
          65 70 75 80
          Lys Trp Glu Arg His Ile Glu His Tyr Gln Glu Glu Pro Asn Lys His
                          85 90 95
          Gly Ile Tyr Leu Gln Asn Ile Leu Pro Pro Leu Arg Glu Asn Pro Arg
                      100 105 110
          Trp Thr Leu Pro Leu Val Cys Arg Ile Ser Gly Tyr Phe Gln Ser Tyr
                  115 120 125
          His Tyr Val Ser Thr Asp Phe Arg Arg Arg Val Gly Lys Phe Leu Asn
              130 135 140
          Leu His Ala Thr Ser Ala Asp Leu Ala Arg Glu Tyr Ala Leu Met Ile
          145 150 155 160
          Gln Pro Glu Asp Val Ala Ile His Leu Arg Arg Gly Asp Phe Val Ala
                          165 170 175
          Leu Gln His Thr Gly Val Gln Leu Phe Gly Ala Glu His Tyr Ala Lys
                      180 185 190
          Gly Leu Ala Leu Gln Ala Gln Gln Gln Pro Ile Gly Arg Val Phe Val
                  195 200 205
          Phe Ser Asp Asp Phe Asp Ala Ile Gln Glu Glu Leu Gly Leu Leu Ala
              210 215 220
          Asp Asn Tyr Gln Leu Val Leu Val Lys Gly Leu Thr Pro Leu Gln Asp
          225 230 235 240
          Leu Phe Leu Leu Thr Cys Phe Arg Arg Tyr Val Leu Ala Asn Ser Thr
                          245 250 255
          Phe Ser Trp Trp Gly Ala Leu Cys Ser Lys Tyr Gly Asn Glu Val Lys
                      260 265 270
          Val Val Val Pro Lys Lys Pro Leu Leu Ile Ser Tyr Pro Glu Asp Ser
                  275 280 285
          Tyr Phe Pro Pro Ser Trp Glu Gln Ile
              290 295
           <![CDATA[ <210> 26]]>
           <![CDATA[ <211> 281]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Campylobacter mucosa]]>
           <![CDATA[ <400> 26]]>
          Met Ile Ile Val Lys Ile Val Gly Gly Leu Gly Asn Gln Met Phe Cys
          1 5 10 15
          Tyr Ala Tyr Ala Lys Met Leu Ser Ile Gly Gly Tyr Asn Val Lys Met
                      20 25 30
          Asp Thr Ser Val Tyr Lys Thr Tyr Lys Leu His Gly Gly Tyr Gln Leu
                  35 40 45
          Asp Lys Tyr Asn Ile Asp Leu Ala Thr Leu Ser Gly Asn Lys Asn Ala
              50 55 60
          Lys Phe Tyr Ser Asn Gly Val Val Phe Lys Ile Leu Lys Lys Phe Gly
          65 70 75 80
          Leu Leu Lys Ile Lys Arg Glu Glu Ser Leu Leu Phe Asp Glu Ser Phe
                          85 90 95
          Lys Thr Pro Asn Asp Lys Val Tyr Ile Glu Gly Tyr Phe Gln Ser Glu
                      100 105 110
          Lys Tyr Phe Cys Asp Ile Lys Asp Ile Leu Leu Asn Gln Phe Val Leu
                  115 120 125
          Lys Thr Thr Leu Ser Asn Tyr Ala Ile Ser Ile Lys Asp Lys Ile Thr
              130 135 140
          Gln Asn Ser Cys Ser Ile His Ile Arg Arg Gly Asp Tyr Leu Leu Gly
          145 150 155 160
          Lys Asn Ile Asn Tyr His Gly Val Cys Asp Leu Lys Tyr Tyr Gln Asp
                          165 170 175
          Ala Thr Asp Phe Leu Glu Gln Lys Cys Gly Asp Leu Thr Tyr Phe Ile
                      180 185 190
          Phe Ser Asp Asp Ile Glu Trp Ala Lys Gly Asn Leu Lys Leu Asn Asn
                  195 200 205
          Ala Phe Tyr Val Asp Ser Ser Asp Arg Leu Pro His Glu Asp Ile Tyr
              210 215 220
          Leu Met Ser Leu Cys Gln Asn Asn Ile Ile Ala Asn Ser Thr Phe Ser
          225 230 235 240
          Trp Trp Gly Ala Tyr Leu Asn Gln Asn Gln Asn Lys Ile Val Ile Ala
                          245 250 255
          Pro Lys Ile Trp Phe Ser Asn Ala Lys Leu Gln Ser Gln Thr Ser Asp
                      260 265 270
          Leu Ile Pro Asn Glu Trp Ile Arg Val
                  275 280
           <![CDATA[ <210> 27]]>
           <![CDATA[ <211> 283]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Geobacter kyunghee]]>
           <![CDATA[ <400> 27]]>
          Met Ile Tyr Val Val Phe Ser Asp Arg Leu Gly Asn Asn Leu Phe Gln
          1 5 10 15
          Met Ala Ala Ala Leu Ser Leu Ser Glu Lys Ile Thr Ile Cys Val Pro
                      20 25 30
          Leu Gln Glu Glu Tyr Ser Ala Thr Leu Lys Tyr Lys Asp Thr Phe Phe
                  35 40 45
          Lys Gly Phe Asp Ile Leu Asn Tyr Ile Pro Asp Gly Ile Pro Val Tyr
              50 55 60
          Glu Glu Pro Phe Tyr His Tyr Asn Ser Val Pro Tyr Thr Glu Gly Thr
          65 70 75 80
          Asp Leu Ile Ile Lys Gly Tyr Phe Gln Ser His Arg Tyr Phe Asp Arg
                          85 90 95
          Ser Leu Val Leu Lys Gln Tyr Ser Ile Asp Asp Asn Thr Ile Phe Phe
                      100 105 110
          Ile Glu Lys Lys Tyr Pro Glu Ile Leu Lys Gly Gly Tyr Thr Ser Ile
                  115 120 125
          His Ile Arg Arg Gly Asp Tyr Leu Lys Met Leu Tyr Lys His Pro Phe
              130 135 140
          Cys Gly Leu Lys Tyr Tyr Lys Lys Ala Ile Asp Leu Ile Gly Lys Asn
          145 150 155 160
          Glu Asn Phe Ile Ile Val Ser Asp Asp Ile Thr Trp Cys Lys Asn Asn
                          165 170 175
          Ile Lys Leu Lys Asn Val Ile Phe Ala Glu Asn Thr Ser Pro Ile Ile
                      180 185 190
          Asp Leu Tyr Ile Gln Ser Tyr Cys Glu Asn Asn Ile Leu Ser Asn Ser
                  195 200 205
          Ser Phe Ser Trp Trp Gly Ala Tyr Leu Asn Lys His Ile Asn Lys Lys
              210 215 220
          Val Ile Val Pro Ser Leu Trp Phe Gly Tyr Lys Ala Asn Phe Ser Thr
          225 230 235 240
          Lys Asp Leu Leu Pro Arg Asp Tyr Ile Val Ile Ile Asn Lys Tyr Ser
                          245 250 255
          Phe Val Gly Tyr Leu Lys Ser Val Met Gln Leu Val Lys Asn Lys Ile
                      260 265 270
          Phe Phe Leu Leu Asn Asn Lys Asp Val Gln Asn
                  275 280
           <![CDATA[ <210> 28]]>
           <![CDATA[ <211> 286]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Autotrophic Neospira spirochetes]]>
           <![CDATA[ <400> 28]]>
          Met Ile Val Val Arg Leu Ile Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Ser Ala Leu Ala Leu Ala Lys Arg Thr Gly Ser Asp Leu Ile
                      20 25 30
          Leu Asp Leu Ser Ala Phe Lys His Tyr Ser Leu Arg Asp Tyr Gly Leu
                  35 40 45
          Gln Gln Trp Lys Ile Ser Ala Arg Ile Ala Ser Ser Val Glu Leu Arg
              50 55 60
          Lys Phe Pro Arg Trp Gly Ala Arg Val Ala Arg Tyr Thr Gln Lys Leu
          65 70 75 80
          Gly Ile Lys Thr Arg Phe Tyr Ser Glu Leu Gly Leu Gly Phe Asp Pro
                          85 90 95
          Ala Phe Met Lys Leu His Ala Pro Val Tyr Leu Asn Gly Tyr Phe Gln
                      100 105 110
          Ser Glu Lys Tyr Phe Leu Asn Ile Arg Ser Ile Leu Leu Glu Glu Phe
                  115 120 125
          Gln Pro Ser Asn Pro Leu Leu Ala Gln Asn Ser Arg His Ala Ser Asp
              130 135 140
          Ala Ile Glu Ser Asn Ser Val Ala Ile His Ile Arg Arg Gly Asp Tyr
          145 150 155 160
          Val Ser Asn Thr His Asn Leu Ser Ile His Gly Val Cys Ser Leu Gly
                          165 170 175
          Tyr Tyr Glu Arg Ala Ile Asn Ile Ile Arg Ser Lys Ile Gly Pro Ala
                      180 185 190
          Asn Phe Phe Val Phe Ser Asp Asp Met Glu Trp Val Arg Glu Asn Ile
                  195 200 205
          Arg Ile Pro Asp Glu Val Val Phe Val Glu Gly Asn Asn Ala Arg Pro
              210 215 220
          Glu Ala Asp Ile His Leu Met Ser Leu Cys Lys His Asn Ile Cys Ala
          225 230 235 240
          Asn Ser Ser Phe Ser Trp Trp Gly Ala Trp Leu Asn Arg Asn Pro Glu
                          245 250 255
          Lys Ile Val Ile Ala Pro Asp Pro Trp Phe Val Ser Lys Ala Leu Asp
                      260 265 270
          Ser Asn His Leu Ile Pro Gln Asp Trp Phe Gln Val Ser Ser
                  275 280 285
           <![CDATA[ <210> 29]]>
           <![CDATA[ <211> 294]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Microbacterium trichothecenes]]>
           <![CDATA[ <400> 29]]>
          Met Thr Val Arg Asp Gly Val Cys Ala Tyr Val Met Gly Gly Leu Gly
          1 5 10 15
          Asn Gln Leu Phe Ile Leu Ala Ala Ala Trp Glu Gln Ser Arg Arg Leu
                      20 25 30
          Asn Val Pro Leu Tyr Leu Asp Arg Ser His Phe Ala Val Gly Gly Thr
                  35 40 45
          Phe Ala Tyr Gly Leu Asp Ala Val Pro Asn Pro Gly His Val Leu Ser
              50 55 60
          Pro Thr Glu Ser Pro Trp Arg Ser Val Arg Ile Asn Arg Glu Arg Val
          65 70 75 80
          Leu Pro Leu Pro Arg Arg Pro Trp Gly Arg Val Tyr Leu Glu Arg Asp
                          85 90 95
          Gly Asp Thr Tyr Ser Pro Ala Ile Asn Gly Ile Arg Pro Gly Thr Thr
                      100 105 110
          Leu Val Gly Tyr Phe Gln Ser Pro Arg Tyr Phe Pro Thr Val Arg Asp
                  115 120 125
          Glu Leu Ala Ser Ala Met Leu Lys Thr Glu Glu Thr Ala Glu Glu Thr
              130 135 140
          Ala Arg Ile Glu Ser Met Leu Ala Ala Pro Ala Ile Thr Leu His Leu
          145 150 155 160
          Arg Arg Gly Asp Tyr Leu Ala Val Ser Ala Asp Arg Gln Phe Ile Ala
                          165 170 175
          Ser Val Asp Tyr Ala Val Arg Gly Val Arg His Leu Arg Ser Met Gly
                      180 185 190
          Leu Asp His Pro Val Arg Val Phe Ser Asp Ser Val Asp Leu Val Lys
                  195 200 205
          Asp Glu Leu Arg Gly Val Glu Gly Asp Phe Glu Phe Val Glu Asp Asp
              210 215 220
          Gly Val Leu Gly Thr Trp Ala Thr Leu Lys Ala Met Ser Ala Gly Thr
          225 230 235 240
          Ala Met Ile Met Ser Asn Ser Ser Phe Ser Trp Trp Gly Ala Glu Leu
                          245 250 255
          Met Arg Gly Arg Leu Gly Pro Gly Val Pro Val Ile Ala Pro Arg Pro
                      260 265 270
          Trp Thr Gln Thr Gly Thr Ala Lys Ala Asp Leu Leu Glu Pro Ala Trp
                  275 280 285
          Val Thr Leu Asp Ala Arg
              290
           <![CDATA[ <210> 30]]>
           <![CDATA[ <211> 272]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Candidate protist cell surface symbionts]]>
           <![CDATA[ <400> 30]]>
          Met Asn Val Val Gln Leu Gln Gly Gly Leu Ala Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Phe Gly Gln Ala Leu Gly Glu Asp Thr Leu Tyr Asp Val Ser
                      20 25 30
          Trp Phe Glu Arg Ala Lys Ser Glu Arg Thr Thr Ser Arg Pro Phe Cys
                  35 40 45
          Leu Asp Val Phe Asn Ile Pro Ala Thr Val Phe Val Asn Arg Arg Pro
              50 55 60
          Pro Leu Leu Ala Arg Leu Val Gly Lys Tyr Lys Val Val Arg Glu Arg
          65 70 75 80
          Asn Trp Asp Glu Phe Gln Pro Glu Phe Leu Asn Leu Gly Gly Arg Gly
                          85 90 95
          Thr Asn Tyr Phe Lys Gly Tyr Phe Gln Asn Glu Arg Tyr Leu Asn Cys
                      100 105 110
          Val Lys Asp Lys Ile Ser Ala Asp Phe Ser Leu Lys Asn Thr Met Asn
                  115 120 125
          Ser Asp Asn Leu Glu Leu Leu Lys Gln Ile Arg Asn Cys Asn Ser Val
              130 135 140
          Ser Leu His Ile Arg Arg Gly Asp Tyr Val Lys Gln Pro Asp Tyr His
          145 150 155 160
          Pro Leu Cys Gly Ile Glu Tyr Tyr Gln Ala Ala Met Lys Met Ile Glu
                          165 170 175
          Gly His Tyr Phe Leu Phe Ser Asp Asp Ile Glu Trp Cys Lys Ala Asn
                      180 185 190
          Ala Lys Thr Asp Met Pro Met Thr Ile Val Asp Ile Asn Ser Ala Glu
                  195 200 205
          Thr Gly Tyr Trp Asp Met Glu Leu Met Lys Asn Cys Lys His Asn Ile
              210 215 220
          Ile Ala Asn Ser Ser Phe Ser Trp Met Ala Ala Tyr Leu Asn Pro Asn
          225 230 235 240
          Pro Asp Lys Met Val Ile Ala Pro Lys Asp Trp Asn Lys His Asn Gly
                          245 250 255
          Glu Ala Ile Ser Asn Arg Gly Leu Ser Glu Lys Tyr Ile Gln Ile Lys
                      260 265 270
           <![CDATA[ <210> 31]]>
           <![CDATA[ <211> 275]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Geobacter sp.]]>
           <![CDATA[ <400> 31]]>
          Met Ile Gly Ile Ser Ile Gln Cys Arg Leu Gly Asn Gln Leu Phe Gln
          1 5 10 15
          Tyr Ala Phe Ile Ile Ser Leu Ser Lys Thr Leu Asn Ser Lys Tyr Phe
                      20 25 30
          Ile Ser Glu Lys Ile Glu Arg Phe Thr Leu Pro Asp Tyr Phe Glu Leu
                  35 40 45
          Pro His Tyr Asn Pro Asn Leu Asn Leu Leu Tyr Lys Ile Ile Leu Lys
              50 55 60
          Ile Lys Lys Gly Tyr Leu Arg Lys Pro Leu Gln Asn Tyr Ser Leu Thr
          65 70 75 80
          Arg Asn Asn Ile Ala Arg Asp Asn Glu Ile Tyr Val Gly Tyr Phe Gln
                          85 90 95
          Ser Glu Thr Phe Phe Asn Gln Leu Lys Asn Asp Ile Asp Ser Phe Ile
                      100 105 110
          Lys Val Arg Lys Glu Tyr Lys Gln Val Phe Ser Asn Gln Tyr Gly Asn
                  115 120 125
          Phe Phe Ser Glu His Lys Thr Ile Ala Ile His Leu Arg Arg Gly Asp
              130 135 140
          Tyr Leu Asn Leu Asp Asn Trp Trp Leu Glu Asn Leu Gly Ser Asn Asn
          145 150 155 160
          Leu Thr Leu Pro Leu Asp Tyr Tyr Lys Asn Cys Phe Ala Ala Ile Glu
                          165 170 175
          Asn Leu His His Tyr Lys Leu Met Phe Ile Ser Asp Asp Ile Glu Phe
                      180 185 190
          Val Lys Ser Glu Phe Gly Tyr Leu Glu Asn Ala Glu Phe His Asn Asn
                  195 200 205
          Glu Leu Ile Ile Asp Phe Gln Ile Met Met Asn Ala Asp Ile Cys Ile
              210 215 220
          Val Ser Asn Ser Ser Phe Ala Trp Trp Ala Ala Tyr Leu Asn Pro Lys
          225 230 235 240
          Ala Thr Lys Lys Ile Leu Cys Pro Lys Tyr Trp Leu Gly Phe Asn Ile
                          245 250 255
          Lys Lys Glu Tyr Pro Lys Asn Ile Ile Pro Lys Asp Trp Lys Gln Ile
                      260 265 270
          Ile Ala Lys
                  275
           <![CDATA[ <210> 32]]>
           <![CDATA[ <211> 288]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Aureobacter sp.]]>
           <![CDATA[ <400> 32]]>
          Met Thr Ile Thr Lys Leu Gln Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Val Ala Arg Ser Arg Cys Val Asp Glu Lys Ile Tyr Leu Asp
                      20 25 30
          Phe Ser Phe Leu Gln Lys Asn Asn Val Ser Thr Glu His Phe Thr Lys
                  35 40 45
          Arg Asp Phe Glu Leu Gly Val Phe Gly Ile Gln Tyr Lys Glu Phe Ser
              50 55 60
          Asp Arg Glu Arg Lys Ile Cys Phe Gly Thr Ser Thr Lys Asp Lys Ile
          65 70 75 80
          Leu Arg Lys Leu Phe Tyr Gly Arg Thr Glu Thr Val His Gln Ile Glu
                          85 90 95
          Asn Glu Phe Val Ile Ile Pro Ala Ala Gln Asn Ile Tyr Phe Asp Gly
                      100 105 110
          Tyr Phe Gln Ser Glu Lys Tyr Phe Ala Ser Ile Arg Asn Gln Leu Leu
                  115 120 125
          Thr Glu Phe Thr Phe Pro Glu Leu Asp Gly Ala Asn Ile Arg Ile Leu
              130 135 140
          Asp Gln Ile Asn Ser Val Gln Asn Ser Val Ser Ile His Ile Arg Arg
          145 150 155 160
          Gly Asp Tyr Leu Lys Pro Glu Val Leu Lys Tyr His Gly Ser Leu Gly
                          165 170 175
          Glu Asn Tyr Tyr Lys Ala Gly Leu Glu Leu Leu Lys Ala Lys Phe Pro
                      180 185 190
          Asn Leu Tyr Tyr Phe Val Phe Ser Asp Asp Ile Ser Phe Ala Lys Thr
                  195 200 205
          Leu Phe Lys Asp Leu Glu Asn Thr Tyr Phe Val Asn Ile Asn Ser Gly
              210 215 220
          Lys Asp Ser Trp Lys Asp Met Phe Leu Met Ala Gln Cys Arg His His
          225 230 235 240
          Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala Trp Leu Ser Gln
                          245 250 255
          Lys Gln Gly Ile Asn Ile Ala Pro Glu Asn Trp Phe Asn Lys Glu Val
                      260 265 270
          Ala Asn Phe Asp Ile Asn Asn Ile Val Pro Ser Asp Trp Ile Thr Ile
                  275 280 285
           <![CDATA[ <210> 33]]>
           <![CDATA[ <211> 283]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Microbacterium sp.]]>
           <![CDATA[ <400> 33]]>
          Met Gly Gly Leu Gly Asn Gln Ile Phe Ile Leu Ala Ala Ala Trp Glu
          1 5 10 15
          Gln Ala Arg Arg Leu Asp Val Pro Leu Tyr Leu Asp Ala Ser His Phe
                      20 25 30
          Ser Val Gly Gly Thr Phe Arg Tyr Gly Leu Asp Ala Ile Asp His Pro
                  35 40 45
          Gly Arg Leu Leu Gly Pro Lys Glu Ser Pro Trp Arg Ser Val Arg Ile
              50 55 60
          Asn Arg Glu Arg Val Leu Pro Phe Pro Arg Arg Pro Trp Gly His Ile
          65 70 75 80
          Phe Leu Glu Arg Asp Gly Asp Arg Tyr Gln Pro Ala Ile Glu Gln Ile
                          85 90 95
          Thr Ser Gly Thr Thr Leu Val Gly Tyr Phe Gln Ser Pro Arg Tyr Phe
                      100 105 110
          Pro Asn Val Arg Asp Gln Leu Ile Ala Ser Met Leu Asp Thr Ala Asp
                  115 120 125
          Thr Glu Ala Glu Ala Ser Arg Leu Arg Glu Tyr Asp Arg Thr Pro Ala
              130 135 140
          Ile Ser Leu His Leu Arg Arg Gly Asp Tyr Leu Ala Val Ser Ala Asp
          145 150 155 160
          Arg Gln Phe Ile Ala Ser Val Ala Tyr Ala Val Arg Ser Val Lys Leu
                          165 170 175
          Leu Arg Ala Met Gly Ile Asp Leu Pro Val Arg Val Phe Ser Asp Ser
                      180 185 190
          Val Asp Leu Val Lys Asp Glu Leu Arg Gly Val Glu Gly Asp Phe Asp
                  195 200 205
          Phe Val Glu Asp Asp Gly Ala Leu Gly Thr Trp Ala Thr Leu Lys Ala
              210 215 220
          Met Ser Ala Gly His Ala Met Ile Met Ser Asn Ser Ser Phe Ser Trp
          225 230 235 240
          Trp Ala Ala Glu Leu Met His Arg Arg Ser Gly Gly Arg Ala Pro Val
                          245 250 255
          Ile Ala Pro Arg Pro Trp Thr Gln Thr Gly Thr Ala Lys Ser Asp Leu
                      260 265 270
          Leu His Ala Asp Trp Ile Thr Leu Asp Ala Arg
                  275 280
           <![CDATA[ <210> 34]]>
           <![CDATA[ <211> 264]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Water Bears]]>
           <![CDATA[ <400> 34]]>
          Met Phe Ile Tyr Ala Ser Leu Trp Gly Ile Ala Lys Lys Asn Gly Leu
          1 5 10 15
          Thr Pro Thr Ile Pro Phe Thr Lys Leu His Asp Ala Phe His Leu Asp
                      20 25 30
          Leu Ser Pro Glu Gln Phe Phe Asp Leu Ser Asp Cys Asp Asn Tyr Val
                  35 40 45
          Phe Gly Met Asp Glu Cys Cys Leu Tyr Asp Glu Arg Thr Glu Gln Ile
              50 55 60
          Phe Gln Arg Ala Val Gly Arg Asn Val Ser Leu Trp Gly Tyr Phe Gln
          65 70 75 80
          Ser Ser Arg Tyr Phe His Pro Glu His Glu Thr Asp Ile Arg Arg Gln
                          85 90 95
          Phe Ala Phe Lys Glu Asn Ile Asp Val Arg Ala Asn Asp Ile Ile Lys
                      100 105 110
          Gly Phe Arg Leu Arg Ser Gly Asp Ser Met Gln Lys Arg Pro Ser Val
                  115 120 125
          Gly Ile His Ile Arg Arg Gly Asp Val Leu Val Lys Ser Phe Trp Arg
              130 135 140
          Asn Phe Gly Phe Thr Val Ala Thr Arg Glu Tyr Phe Arg His Ala Met
          145 150 155 160
          Glu Tyr Leu Lys Asn Lys Gly Val Ile Asp Pro Ile Phe Leu Val Cys
                          165 170 175
          Thr Asn Asp Val Glu Trp Ser Arg Gln Asn Leu Ala Gly Phe Asp Val
                      180 185 190
          His Phe Val Glu Asn Gln Thr Ser His Val Asp Met Ala Ile Leu Thr
                  195 200 205
          His Met Asn His Leu Ile Leu Ser Ala Gly Thr Phe Ser Phe Phe Ala
              210 215 220
          Gly Tyr Leu Ser Ser Ala Gln His Ile Ile Tyr Tyr Lys Gly Trp Pro
          225 230 235 240
          Arg Pro Gly Ser Lys Tyr Ala Gly Met Met Asp Leu Ser Ser Phe Trp
                          245 250 255
          Leu Pro Glu Trp Ile Ala Met Glu
                      260
           <![CDATA[ <210> 35]]>
           <![CDATA[ <211> 288]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Candidate species of Wolffia bacteria]]>
           <![CDATA[ <400> 35]]>
          Met Ile Ile Ile Lys Leu Lys Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Cys Ala Lys His Leu Ala Glu Arg Asn Gly Asp Val Leu Lys
                      20 25 30
          Leu Asp Ile Ser Leu Tyr Gln Pro Gly Asn Ala Pro Ala Gly Asp Thr
                  35 40 45
          Ala Arg Ser Tyr Ala Leu Gly Ala Phe Ser Ile Ser Ala Gln Val Ala
              50 55 60
          Ser Lys Glu Glu Val Arg Arg Val Arg Gly Leu Leu Trp His Ile Met
          65 70 75 80
          Ala Leu Ala Met Lys Val Ile Asn Arg Val Arg Pro Ile Ser Ser Tyr
                          85 90 95
          Met Phe Asp Pro Gln Val Leu Glu Arg Arg Gly Asn Val Tyr Leu Asp
                      100 105 110
          Gly Tyr Phe Gln Ser Glu Arg Tyr Phe Lys Asp Ile Lys Ala Cys Ile
                  115 120 125
          Arg Asn Glu Phe Arg Leu Arg Glu Pro Met Gly Glu Gln Ala Gln Ala
              130 135 140
          Met Leu Arg Asp Ile Asp Gly Ser Asp Ser Val Ser Leu His Ile Arg
          145 150 155 160
          Arg Gly Asp Tyr Val Thr Asn Lys Asn Ala Asn Ala Phe His Gly Thr
                          165 170 175
          Cys Pro Pro Glu Tyr Tyr His Ala Ala Ile Lys Glu Ile Gly Lys His
                      180 185 190
          Val His Ser Pro Lys Phe Phe Ile Phe Ser Asp Asp Ile Glu Trp Ala
                  195 200 205
          Lys Ala Asp Val Gly Met Pro Ala His Ala Thr Tyr Val Ser Arg Asp
              210 215 220
          Gly Ile Ala Asp Tyr Glu Glu Leu Leu Leu Met Ser Lys Cys Lys His
          225 230 235 240
          Asn Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala Trp Leu Asn
                          245 250 255
          Ala Asn Ser Asn Lys Ile Val Val Ala Pro Lys Arg Trp Ile Asn Asp
                      260 265 270
          Glu Arg Val Asp Thr Ser Asp Ala Val Pro Val Glu Trp Val Ser Met
                  275 280 285
           <![CDATA[ <210> 36]]>
           <![CDATA[ <211> 272]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Candidate species of Yanofsky bacteria]]>
           <![CDATA[ <400> 36]]>
          Met Ile Thr Phe Arg Lys Leu Gly Leu Leu Gly Arg Phe Gly Asn Gln
          1 5 10 15
          Leu Phe Gln Tyr Ala Gly Ala Arg Met Tyr Ala Asp Ile Asn Gly Phe
                      20 25 30
          Lys Ser Ala Phe Pro Gly Trp Val Gly Asn Lys Ile Phe Glu Asn Ile
                  35 40 45
          Ile Ser Tyr Asn Asn Arg Glu Tyr Leu Leu Ser Arg Phe Leu Pro Thr
              50 55 60
          Arg Gln Leu Asn Asp Phe Leu Ser Tyr Thr Arg Thr Asp Lys Ile Lys
          65 70 75 80
          Tyr Met Leu Arg Leu Gln Lys Gln Leu Pro Gln Thr Ile Ala Leu Asn
                          85 90 95
          Lys Leu Tyr Ala His Pro Glu Asp Asn Ile Asn Phe Leu Gly Tyr Phe
                      100 105 110
          Gln Asn Glu Leu Ser Leu Lys Ile Leu Lys Glu Arg Lys Asn Tyr Val
                  115 120 125
          Leu Gly Trp Phe Val Phe Lys Lys Asp Ile Ala Asp Glu Phe Lys Lys
              130 135 140
          Val Thr Gln Asn Tyr Lys Pro Trp Thr Gly Ile His Ile Arg Arg Gly
          145 150 155 160
          Asp Phe Val Lys Arg Gly Leu Ser Leu Pro Val Phe Leu Tyr Lys Asp
                          165 170 175
          Phe Leu Lys Ser Val Asp Lys Lys Asn Ile Tyr Ile Ser Cys Asp Asp
                      180 185 190
          Pro Glu Thr Ile Ser Glu Phe Lys Asp Tyr Ser Leu Ile Arg Pro Ala
                  195 200 205
          Asn Pro Leu Pro Glu Ile Pro Asn Phe Ile Phe Asp Phe Trp Met Leu
              210 215 220
          Lys Glu Ser Glu Met Val Leu Gly Cys Gly Ser Thr Phe Ser Trp Trp
          225 230 235 240
          Ala Ala Tyr Leu Gly Asn Arg Asn Asn Tyr Phe Ser Pro Pro Leu Thr
                          245 250 255
          His Leu Trp Pro Lys Gly Tyr Lys Pro Thr Leu Gly Lys Met Asp Ile
                      260 265 270
           <![CDATA[ <210> 37]]>
           <![CDATA[ <211> 276]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Spirochetes]]>
           <![CDATA[ <400> 37]]>
          Met Ile Gly Ile Lys Ile Gly Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Phe Gly Tyr Val Val Ala Lys Lys Phe Ser Thr Ser Phe Tyr
                      20 25 30
          Leu Glu Lys Arg Asn Gln Phe Lys Leu Asp Lys Tyr Phe Ser Leu Arg
                  35 40 45
          Ile Leu Glu Asn Ser Ile Asn Ser Leu Ile Lys Pro Leu Tyr Lys Met
              50 55 60
          Gln Thr Lys Leu Gly Lys Ile Lys Met Tyr Asp Trp Glu Asp Phe Gly
          65 70 75 80
          Thr Lys Thr Pro Glu Ser Phe Ile Ser Gln Ile Lys Asp Asn Ile Tyr
                          85 90 95
          Tyr Cys Gly Tyr Phe Gln Ser Glu Lys Tyr Phe Glu Gly Phe Tyr Lys
                      100 105 110
          Asp Leu Gln Lys Ile Phe Lys Ile Lys Lys Glu Tyr Ile Glu Gln Phe
                  115 120 125
          Asn Ser Lys Tyr Glu Lys Phe Tyr Lys Glu Asn Lys Val Ile Ala Ile
              130 135 140
          His Ile Arg Arg Gly Asp Tyr Ile Asn Phe Gly Asp Asp Ser Leu Gly
          145 150 155 160
          Gly Lys Asn Met Thr Leu Pro Ile Ser Tyr Tyr Thr Asn Cys Leu Lys
                          165 170 175
          Gln Ile Lys Asp Ile Ser Ser Tyr Lys Ile Ile Phe Ile Ser Asp Asp
                      180 185 190
          Ile Glu Phe Val Lys Lys Glu Phe Gly Gln Lys Glu Asn Tyr Phe Phe
                  195 200 205
          Glu Ser Asn Asn Glu Ile Ile Asp Phe Gln Val Ile Leu Asn Ala Asp
              210 215 220
          Lys Val Ile Met Ser Asn Ser Ser Phe Ser Trp Trp Ala Ser Tyr Leu
          225 230 235 240
          Asn Ile Lys Ala Thr Asp Ile Tyr Cys Pro Asn Tyr Trp Leu Gly Phe
                          245 250 255
          Lys Ile Gly Thr Glu Asn Pro Ile Gly Ile Val Cys Lys Lys Trp Thr
                      260 265 270
          Arg Leu Asp Ile
                  275
           <![CDATA[ <210> 38]]>
           <![CDATA[ <211> 296]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Geobacter agrobacterium]]>
           <![CDATA[ <400> 38]]>
          Met Glu Thr Glu Ile Pro Val Thr Gln Asp Leu Asp Gln Asn Lys Gln
          1 5 10 15
          Val Tyr Leu Ile Thr Met Val Gly Ile Ser Ile Gln Cys Arg Leu Gly
                      20 25 30
          Asn Gln Leu Phe Gln Tyr Ala Phe Ile Thr Ala Leu Ser Lys Lys Leu
                  35 40 45
          Asn Thr Lys Tyr Phe Ile Ser Glu Lys Ile Glu Arg Phe Thr Leu Pro
              50 55 60
          Asp Tyr Phe Glu Leu Pro Gln Tyr Ser Ser Lys Leu Asn Phe Leu Arg
          65 70 75 80
          Lys Leu Ile Leu Lys Ile Lys Lys Gly Tyr Thr Leu Lys Pro Leu Gln
                          85 90 95
          His Tyr Glu Ile Thr Gln Asn Thr Pro Ala Lys Asp Asn Glu Ile Tyr
                      100 105 110
          Val Gly Tyr Phe Gln Ser Glu Ser Phe Phe Glu Gly Thr Lys Asp Asn
                  115 120 125
          Ile Gly Glu Leu Ile Lys Val Arg Pro Glu His Lys Asn Ala Phe Asn
              130 135 140
          Asn Gln Phe Glu Lys Thr Phe Ser Gln Gln Lys Thr Ile Ala Ile His
          145 150 155 160
          Leu Arg Arg Gly Asp Tyr Leu Asn Leu Asp Asp Trp Trp Leu Glu Asn
                          165 170 175
          Leu Gly Gly Asn Asn Leu Thr Leu Pro Ile Thr Tyr Tyr Glu Lys Cys
                      180 185 190
          Phe Ala Ala Ile Lys Asn Leu Asn Asn Tyr Arg Leu Leu Phe Ile Ser
                  195 200 205
          Asp Asp Ile Ala Phe Ala Arg Leu Asn Phe Gly His Leu Glu Asn Ala
              210 215 220
          Glu Phe His Asn Asn Glu Leu Ile Ile Asp Phe Gln Ile Met Met Asn
          225 230 235 240
          Ala Asp Ile Cys Ile Val Ser Asn Ser Ser Phe Ala Trp Trp Ala Ala
                          245 250 255
          Tyr Leu Asn Ala Lys Ser Thr Lys Lys Ile Phe Cys Pro Glu Tyr Trp
                      260 265 270
          Leu Gly Phe Asn Ile Asn Thr Glu Tyr Pro Lys Asn Ile Ile Pro Asp
                  275 280 285
          Ser Trp Glu Gln Ile Ala Val Lys
              290 295
           <![CDATA[ <210> 39]]>
           <![CDATA[ <211> 304]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Bacteroides KHT7]]>
           <![CDATA[ <400> 39]]>
          Met Glu Asn Asn Ile Leu Gln Thr Thr Leu Leu Gly Arg Leu Gly Asn
          1 5 10 15
          Gln Leu Phe Ile Tyr Ala Ala Thr Arg Val Met Ala Leu Glu Ser Gly
                      20 25 30
          Lys Lys Leu Lys Leu Thr Cys Asp Ala Met Glu Arg Phe Arg Ala Thr
                  35 40 45
          Tyr Arg Leu Asp Cys Phe Asn Ile Pro Gln Asp Gln Val Ser Leu Val
              50 55 60
          Ser Asp Lys His Leu Ser Ile Lys Gln Lys Ile Gly Tyr Phe Phe Tyr
          65 70 75 80
          Leu Leu Lys Cys Arg His Lys Thr Val Val Gln Ile His Glu Leu Glu
                          85 90 95
          Asn Lys Tyr Gln Lys Phe Tyr His Arg Phe Gly Leu Ile Leu Asn Gln
                      100 105 110
          Glu Gly Tyr Ile Lys Pro Leu Leu Leu Glu Gly Gly Asn Trp Tyr Ala
                  115 120 125
          Leu Gly Tyr Phe Gln Ser Asp Lys Tyr Phe Thr Lys Tyr Glu Asp Leu
              130 135 140
          Ile Arg Lys Glu Leu Thr Phe Lys Thr Asp Met Phe Cys Lys Asp Ser
          145 150 155 160
          Lys Met Leu Gly Ser Lys Leu Arg Ala Asn Lys Asn Ser Val Cys Leu
                          165 170 175
          His Ile Arg Arg Gly Asp Tyr Leu Asn Asp Pro Val Phe Ala Val Cys
                      180 185 190
          Asp Ile Asn Tyr Tyr Asn Lys Ala Ile Asp Lys Leu Leu Glu Leu Ile
                  195 200 205
          Pro Asn Ala Glu Phe Phe Val Phe Ser Asp Ser Ile Asp Glu Val Arg
              210 215 220
          Glu Phe Leu Ala Lys Tyr Cys Asp Asn Tyr Phe His Tyr Ile Asp Val
          225 230 235 240
          Lys Tyr Thr Asp Gln Glu Ser Leu Tyr Leu Gly Ser Cys Cys Asn Asn
                          245 250 255
          Phe Ile Met Thr Asn Ser Thr Phe Ser Trp Trp Met Gln Tyr Leu Ser
                      260 265 270
          Asp Asn Pro Asn Lys Ile Val Leu Ala Pro Asn Arg Trp Tyr Asn Thr
                  275 280 285
          Asn Asn Pro Cys Asp Ile Tyr Gln Asp Asn Trp Met Ile Ile Glu Val
              290 295 300
           <![CDATA[ <210> 40]]>
           <![CDATA[ <211> 292]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Vibrio butyricum sp. TB]]>
           <![CDATA[ <400> 40]]>
          Met Ile Ile Ile Lys Met Gln Gly Gly Leu Gly Asn Gln Leu Phe Leu
          1 5 10 15
          Tyr Gly Leu Tyr Lys Glu Leu Leu Tyr Leu Gly Arg Glu Val Lys Met
                      20 25 30
          Asp Asn Ile Ser Gly Phe Glu Asp Asp Pro Gln Arg Ser Pro Ser Leu
                  35 40 45
          Tyr Lys Leu Gly Ile Ser Tyr Asp Ile Ala Glu Asp Gly Glu Ile Arg
              50 55 60
          Thr Met Arg Asp Ser Tyr Met Asp Pro Leu Ser Arg Ile Arg Arg Lys
          65 70 75 80
          Ile Thr Gly Arg Lys Asn Lys Asp Tyr Tyr Glu Ala Leu Asp Gly Asn
                          85 90 95
          Phe Asp Glu His Val Leu Glu Ala Asp Asp Ile Tyr Leu Asn Gly Tyr
                      100 105 110
          Phe Gln Ser Asp Asp Tyr Phe Lys Asp Asp Glu Val Arg Gln Glu Leu
                  115 120 125
          Ile Ala Asp Ile Cys Lys Asn Lys Asp Leu Tyr Leu Gly Asp Gln Asp
              130 135 140
          Val Thr Asp Ile Arg Asn Ala Ile Arg Ser Ser Asn Ser Val Ser Met
          145 150 155 160
          His Ile Arg Arg Gly Asp Tyr Leu Thr Pro Val Ser Arg Lys Thr Tyr
                          165 170 175
          Gly Gly Ile Cys Thr Asp Asp Tyr Tyr Ala Lys Ala Met Glu Leu Val
                      180 185 190
          Arg Ser Ser Tyr Pro Asp Ala Gln Phe Tyr Ile Phe Ser Asn Asp Ile
                  195 200 205
          Asp Trp Cys Arg Ala Lys Tyr Lys Asp Thr Asp Asn Ile Ser Phe Val
              210 215 220
          Asn Ile Ser Gly Asp Asp Ser Asp Ile Met Glu Phe Phe Leu Met Ser
          225 230 235 240
          Glu Cys Arg His His Ile Leu Ala Asn Ser Ser Phe Ser Trp Trp Ala
                          245 250 255
          Ala Phe Leu Gly Ser Ala Asp Asp Arg Asp Arg Leu Asn Ile Val Pro
                      260 265 270
          Ser Lys Trp Thr Asn Thr Arg Gln Met Lys Asp Ile Tyr Ala Ser Trp
                  275 280 285
          Met Thr Arg Ile
              290
           <![CDATA[ <210> 41]]>
           <![CDATA[ <211> 309]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Vibrio cellolyticus butyricum]]>
           <![CDATA[ <400> 41]]>
          Met Ile Ile Ile Lys Leu Gln Gly Gly Leu Gly Asn Gln Leu Phe Leu
          1 5 10 15
          Tyr Gly Leu Tyr Arg Asn Leu Glu Arg Leu Gly Arg Asp Val Lys Leu
                      20 25 30
          Asp Ile Glu Ser Gly Phe Glu Cys Asp Glu Leu Arg Ser Pro Cys Leu
                  35 40 45
          Asp Gly Met Lys Leu Lys Tyr Glu Val Ala Thr Arg Lys Glu Val Ile
              50 55 60
          Ala Ile Arg Asp Ser Tyr Met Asp Ile Leu Ser Arg Ile Arg Arg Lys
          65 70 75 80
          Ile Thr Gly Arg Lys Thr Phe Asp Tyr Tyr Glu Pro Glu Asp Gly Asn
                          85 90 95
          Phe Asp Pro Lys Val Leu Glu Leu Thr His Ala Tyr Leu Asn Gly Tyr
                      100 105 110
          Phe Gln Ser Glu Lys Tyr Phe Gly Asp Asp Asn Ser Ala Lys Ala Leu
                  115 120 125
          Lys Ile Glu Leu Thr Thr Asn Arg Tyr Asp Ile Leu Ser Ser Ser Asp
              130 135 140
          Leu Val Thr Asp Ile Tyr Asn Asp Ile Arg Lys Ser Glu Ser Val Ser
          145 150 155 160
          Leu His Ile Arg Arg Gly Asp Tyr Leu Thr Pro Gly Ile Ile Glu Thr
                          165 170 175
          Tyr Gly Gly Ile Cys Thr Asp Asp Tyr Tyr Asp Lys Ala Ile Ala Met
                      180 185 190
          Ile Arg Asp Lys His Pro Asp Ala Lys Leu Phe Val Phe Ser Asn Asp
                  195 200 205
          Ile Asp Trp Cys Lys Glu Lys Met Ala Asp Tyr Lys Asn Ile Ser Phe
              210 215 220
          Val Ser Thr Met Pro Val Asn Asn Ile Ile Glu Glu Val Phe Asn Ser
          225 230 235 240
          Ser Gln Ser Ser Lys Ile Thr Asn Asn Tyr Lys Asp Leu Ala Glu Leu
                          245 250 255
          Tyr Leu Met Ser Ala Cys Lys His His Val Leu Ala Asn Ser Ser Ser Tyr
                      260 265 270
          Ser Trp Trp Gly Ala Trp Leu Ser Asp Asn Glu Gly Met Thr Ile Val
                  275 280 285
          Pro Ser Lys Trp Leu Asn Asn Lys Asn Met Thr Asp Ile Tyr Thr Arg
              290 295 300
          Asp Met Ile Leu Ile
          305
           <![CDATA[ <210> 42]]>
           <![CDATA[ <211> 277]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Candidate sire Gassanilacia]]>
           <![CDATA[ <400> 42]]>
          Met Ile Phe Asp Lys Leu Ser Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Ala Gly Tyr Ser Leu Ser Leu Asn Asn Arg Ile Pro Leu Asn
                      20 25 30
          Leu Asp Leu Ser Ser Phe Glu His Lys Lys Thr Gly Ile Thr His Arg
                  35 40 45
          Tyr Phe Leu Leu Asn Lys Phe Asn Ile Asp Arg Asn Ile Leu Ile Asn
              50 55 60
          His Met Glu Lys Ile Ser Gly Tyr Arg Lys Phe Leu Ser Lys Phe Ile
          65 70 75 80
          Thr Lys Phe Phe Gly Glu Asn Phe Tyr Tyr Asn Ile Thr Phe Leu Ser
                          85 90 95
          Ser Lys Tyr Leu Asp Gly Tyr Phe Gln Ser Glu Lys Tyr Phe Lys Asn
                      100 105 110
          Ile Glu Asp Ile Leu Arg Lys Glu Phe Thr Leu Lys Asn Glu Met Ser
                  115 120 125
          Val Val Ala Arg Gln Val Glu Ser Lys Ile Ser Asn Ser Ile Asn Ser
              130 135 140
          Val Ser Leu His Ile Arg Arg Gly Asp Tyr Val Leu Asp Asn Lys Thr
          145 150 155 160
          Asn Ser Tyr His Gly Ile Cys Asp Leu Asp Tyr Tyr Lys Lys Ala Val
                          165 170 175
          Glu Tyr Phe Lys Asn Lys Leu Gly Glu Leu Asn Ile Phe Val Phe Ser
                      180 185 190
          Asp Asp Ile Val Trp Val Lys Glu Asn Leu Arg Phe Glu Asn Leu Tyr
                  195 200 205
          Phe Val Ser Ser Pro Asp Ile Lys Asp Tyr Glu Glu Leu Ile Leu Met
              210 215 220
          Ser Arg Cys Lys His Asn Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp
          225 230 235 240
          Gly Ala Trp Leu Asn Ala Asn Lys Asn Lys Thr Val Ile Thr Pro Lys
                          245 250 255
          Lys Trp Phe Gln Lys Tyr Asn Ile Asn Gln Lys His Ile Val Pro Lys
                      260 265 270
          Ser Trp Ile Arg Leu
                  275
           <![CDATA[ <210> 43]]>
           <![CDATA[ <211> 292]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Vibrio cellolyticus butyricum]]>
           <![CDATA[ <400> 43]]>
          Met Ile Ile Ile Lys Met Gln Gly Gly Leu Gly Asn Gln Leu Phe Leu
          1 5 10 15
          Tyr Gly Leu Tyr Lys Glu Leu Leu Phe Leu Gly Arg Glu Val Lys Met
                      20 25 30
          Asp Asn Asn Ala Gly Phe Ile Glu Asp Pro Leu Arg Asp Pro Val Leu
                  35 40 45
          Ser Lys Leu Gly Ile Ser Tyr Asp Leu Ala Glu Asp Lys Glu Ile Arg
              50 55 60
          Lys Met Arg Asp Ser Tyr Met Asp Pro Leu Ser Arg Ile Arg Arg Lys
          65 70 75 80
          Leu Thr Gly Arg Lys Asn Lys Asp Tyr Tyr Glu Ala Leu Asp Gly Asn
                          85 90 95
          Phe Asp Glu Thr Val Leu Lys Ala Asp Asp Ile Tyr Leu Asn Gly Tyr
                      100 105 110
          Phe Gln Ser Glu Lys Tyr Phe Asn Asp Asp Ser Val Arg Gln Glu Leu
                  115 120 125
          Arg Ala Asp Ile Cys Lys Asn Lys Glu Arg Tyr Met Ser Glu Ser Asp
              130 135 140
          Val Ala Asp Leu Arg Gln Glu Ile Ala Asn Ala Asn Ser Val Ser Met
          145 150 155 160
          His Ile Arg Arg Gly Asp Tyr Leu Thr Pro Val Ser Lys Glu Thr Tyr
                          165 170 175
          Gly Gly Ile Cys Thr Asp Glu Tyr Tyr Glu Lys Ala Met Asp Leu Val
                      180 185 190
          Lys Glu Lys Asn Pro Asp Ala Arg Phe Tyr Ile Phe Ser Asn Asp Ile
                  195 200 205
          Asp Trp Cys Lys Glu Lys Tyr Lys Asp Thr Asp Asn Val Ser Phe Val
              210 215 220
          Ser Leu Thr Gly Asp Asp Ser Asp Ile Lys Glu Phe Phe Leu Met Ser
          225 230 235 240
          Glu Cys Arg His His Ile Leu Ala Asn Ser Ser Phe Ser Trp Trp Ala
                          245 250 255
          Ala Phe Leu Ser Ala Ala Asp Asp Asn Lys Ser Leu Asn Ile Val Pro
                      260 265 270
          Ser Lys Trp Thr Asn Thr Arg Gln Met Lys Asp Ile Tyr Ala Ser Trp
                  275 280 285
          Met Thr Arg Ile
              290
           <![CDATA[ <210> 44]]>
           <![CDATA[ <211> 293]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Turbot aureus]]>
           <![CDATA[ <400> 44]]>
          Met Val Ala Val Glu Leu Ile Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Thr Ala Arg Ala Leu Ala Leu His Arg Asp Glu Pro Leu Ser
                      20 25 30
          Ile Asp Gly Arg Leu Phe Asp Asn Tyr Lys Leu Arg Asn Phe Cys Leu
                  35 40 45
          Ser His Phe Ser Ile Glu Ala Ile Val Val Lys Asn Asp Leu Pro Leu
              50 55 60
          Lys Thr Pro Gly Phe Ser Lys Lys Val Val Asp Gln Ile Leu Lys Lys
          65 70 75 80
          Thr Asn Thr Phe Ile Leu Gln Asn Lys Leu Phe Ser Val Tyr Gln Glu
                          85 90 95
          Lys Asn Leu Leu Phe Asp Asp Ser Leu Phe Arg Asn Gly Lys Lys Asn
                      100 105 110
          Ile Tyr Leu Lys Gly Tyr Phe Gln Ser Glu Lys Tyr Phe Val Lys Tyr
                  115 120 125
          Glu Asp Gln Leu Arg Lys Asp Phe Lys Ile Val Thr Pro Leu Lys Lys
              130 135 140
          Glu Thr Ile Asp Leu Leu Lys Ile Ile Lys Glu Lys Asn Ser Val Ser
          145 150 155 160
          Leu His Ile Arg Arg Gly Asp Tyr Val Ser Asn Pro Thr Val Asn Ala
                          165 170 175
          Ile His Gly Thr Cys Asp Leu Asn Tyr Tyr His Arg Ala Ile Glu Ile
                      180 185 190
          Val Lys Glu Lys Ile Leu His Pro Val Phe Phe Ile Phe Ser Asp Asp
                  195 200 205
          Ile Asp Trp Ala Lys Glu Asn Leu Lys Leu Glu Asn Thr Thr Tyr Phe
              210 215 220
          Val Asp Phe Asn Asp Ala Ser Thr Ser Tyr Glu Asp Leu Lys Leu Met
          225 230 235 240
          Ser Ala Cys Lys His Asn Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp
                          245 250 255
          Gly Ala Trp Leu Asn Thr Asn Lys Ser Lys Thr Val Ile Ala Pro Ser
                      260 265 270
          Lys Trp Phe Asn Thr Asp Val Leu Asn Ser Gln Asp Ile Ile Pro Glu
                  275 280 285
          Ser Trp Met Lys Ile
              290
           <![CDATA[ <210> 45]]>
           <![CDATA[ <211> 261]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Akkermansia sp. 54_46]]>
           <![CDATA[ <400> 45]]>
          Met Asn Met Glu Arg Lys Thr Gly Leu Met Asn Lys Lys Tyr Val Ser
          1 5 10 15
          Pro Cys Phe Leu Pro Gly Met Arg Leu Gly Asn Ile Met Phe Thr Leu
                      20 25 30
          Ala Ala Ala Cys Ala His Ala Arg Thr Val Gly Val Glu Cys Arg Val
                  35 40 45
          Pro Trp Ala Tyr Asn Asp Ala Ser Leu Met Leu Arg Ser Arg Leu Gly
              50 55 60
          Gly Trp Val Leu Pro Ser Thr Pro Cys Gly Thr Asn Glu Pro Ser
          65 70 75 80
          Trp Gln Glu Pro Ser Phe Ser Tyr Cys Pro Val Pro Ser Arg Ile Arg
                          85 90 95
          Thr Gly Gly Leu Arg Gly Tyr Phe Gln Ser Ala Arg Tyr Phe Glu Gly
                      100 105 110
          Gln Glu Ala Phe Ile Arg Ala Leu Phe Ala Pro Leu Thr Ala Glu Lys
                  115 120 125
          Glu Pro Gly Ala Val Gly Ile His Ile Arg Leu Gly Asp Tyr Arg Arg
              130 135 140
          Leu Arg Asp Lys His Arg Ile Leu Asp Pro Gly Phe Leu Arg Arg Ala
          145 150 155 160
          Ala Gly His Leu Ser Ser Gly Lys Asn Arg Leu Val Leu Phe Ser Asp
                          165 170 175
          Glu Pro Asp Glu Ala Ala Glu Met Leu Ala Arg Val Pro Ala Phe Gly
                      180 185 190
          Arg Phe Ala Leu Glu Ile Asp Arg Gly Ala Pro Cys Glu Ser Leu Arg
                  195 200 205
          Arg Met Thr Ala Met Glu Glu Leu Val Met Ser Cys Ser Ser Phe Ser
              210 215 220
          Trp Trp Gly Ala Trp Leu Gly Asn Thr Arg Lys Val Ile Val Pro Arg
          225 230 235 240
          Asp Trp Phe Val Gly Gly Val Glu Asp Tyr Arg Asp Ile Tyr Leu Pro
                          245 250 255
          His Trp Val Thr Leu
                      260
           <![CDATA[ <210> 46]]>
           <![CDATA[ <211> 263]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Shovel-shaped sea bean sprouts]]>
           <![CDATA[ <400> 46]]>
          Met Phe Gln Phe Ala Ser Leu Leu Gly Ile Ala Ala Arg Asn Asn Leu
          1 5 10 15
          Thr Ala Val Ile Pro Tyr Asn Pro Glu Met Glu Arg Ile Phe Lys Ile
                      20 25 30
          Ser Thr Pro Ser Thr Lys Asn Leu Gln Asn Thr Leu Gly Met Tyr Leu
                  35 40 45
          Thr Val Thr Gln Phe Tyr Gly Arg Cys Cys Tyr Tyr Asp Ile Leu Thr
              50 55 60
          Glu Thr Leu Asp Pro Arg His Asn Tyr Leu Leu Tyr Gly Tyr Phe Gln
          65 70 75 80
          Ser Trp Lys Tyr Phe Lys Asn Val Asn Asp Ile Val Arg Gln Glu Phe
                          85 90 95
          Gln Phe Arg Glu Glu Ile Leu Asn Glu Ala Lys Lys Phe His Arg Asp
                      100 105 110
          Val Ser Asn Lys Phe Gln Cys His Glu Asn Asn Ser Cys Thr Phe Val
                  115 120 125
          Gly Ile His Ile Arg Arg Gly Asp Ile Ser Asp Tyr Asp Val Tyr Lys
              130 135 140
          Lys Ala Gly Tyr Thr Thr Pro Ser Glu Gln Tyr Phe Lys Thr Ala Met
          145 150 155 160
          Asn Phe Phe Lys Arg Lys His Lys Asn Lys Val Val Phe Ile Val Cys
                          165 170 175
          Ser Asp Ser Ser Lys Gly Trp Gln Lys Lys Leu Thr Ser Ser Asp Val
                      180 185 190
          Val Phe Ser His Ala Lys Asp Gln Gly Val Asp Leu Ala Ile Leu Ser
                  195 200 205
          Leu Cys Gln His Met Ile Ile Ser Thr Gly Thr Tyr Gly Trp Trp Ala
              210 215 220
          Ala Phe Leu Thr Gly Gly Thr Thr Val Tyr Tyr Arg Asp Trp Pro Lys
          225 230 235 240
          Ala Gly Ser Trp Leu Tyr Asp Gln Val Phe Lys Glu Asp Tyr Phe Val
                          245 250 255
          Arg Gly Trp Ile Pro Met Lys
                      260
           <![CDATA[ <210> 47]]>
           <![CDATA[ <211> 263]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Shovel-shaped sea bean sprouts]]>
           <![CDATA[ <400> 47]]>
          Met Phe Gln Phe Ala Ser Leu Leu Gly Ile Ala Ala Arg Asn Asn Leu
          1 5 10 15
          Thr Ala Val Ile Pro Tyr Asn Pro Glu Met Glu Arg Ile Phe Lys Ile
                      20 25 30
          Ser Thr Pro Ser Thr Lys Asn Leu Gln Asn Thr Leu Gly Met Tyr Leu
                  35 40 45
          Thr Val Thr Gln Phe Tyr Gly Arg Cys Cys Tyr Tyr Asp Ile Leu Thr
              50 55 60
          Glu Thr Leu Asp Pro Arg Tyr Asn Tyr Leu Leu Tyr Gly Tyr Phe Gln
          65 70 75 80
          Ser Trp Lys Tyr Phe Lys Asn Val Asn Asp Ile Val Arg Gln Glu Phe
                          85 90 95
          Gln Phe Arg Glu Glu Ile Leu Asn Glu Ala Lys Lys Phe His Arg Asp
                      100 105 110
          Val Ser Asn Lys Phe Gln Cys His Glu Asn Asn Ser Cys Thr Phe Val
                  115 120 125
          Gly Ile His Ile Arg Arg Gly Asp Ile Ser Asp Tyr Asp Val Tyr Lys
              130 135 140
          Lys Ala Gly Tyr Thr Thr Pro Ser Glu Gln Tyr Phe Lys Thr Ala Met
          145 150 155 160
          Asn Phe Phe Lys Arg Lys His Lys Asn Lys Val Val Phe Ile Val Cys
                          165 170 175
          Ser Asp Ser Ser Lys Gly Trp Gln Lys Lys Leu Thr Ser Ser Asp Val
                      180 185 190
          Val Phe Ser His Ala Lys Asp Gln Gly Val Asp Leu Ala Ile Leu Ser
                  195 200 205
          Leu Cys Gln His Met Ile Ile Ser Thr Gly Thr Tyr Gly Trp Trp Ala
              210 215 220
          Ala Phe Leu Thr Gly Gly Thr Thr Ile Tyr Tyr Arg Asp Trp Pro Lys
          225 230 235 240
          Ala Gly Ser Trp Leu Tyr Asp Gln Val Phe Lys Glu Asp Tyr Phe Val
                          245 250 255
          Arg Gly Trp Ile Pro Met Lys
                      260
           <![CDATA[ <210> 48]]>
           <![CDATA[ <211> 273]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Species of the genus "Ryokoji"]]>
           <![CDATA[ <400> 48]]>
          Met Ile Val Val Gln Leu Ile Gly Gly Leu Gly Asn Gln Leu Phe Gln
          1 5 10 15
          Tyr Ala Ala Ala Lys Ala Phe Ala Leu Glu Thr Glu Gln Lys Leu Tyr
                      20 25 30
          Ile Asp Val Ser Gln Phe Glu Ser Tyr Lys Leu His Asn Tyr Ala Leu
                  35 40 45
          Asn His Phe Asn Ile Ile Ser Asn Val Tyr Asn Lys Pro Asn Arg Tyr
              50 55 60
          Phe Lys Lys Ile Lys Ser Phe Tyr Gln Lys Asn Thr Ser Tyr Lys Glu
          65 70 75 80
          Val Asp Phe Gly Tyr Asn Gln Asp Leu Phe Asn Leu Lys Gly Asp Thr
                          85 90 95
          Ile Phe Leu Asp Gly Tyr Phe Gln Ser Glu Lys Tyr Phe Lys Lys Tyr
                      100 105 110
          Glu Lys Glu Ile Arg Glu Asp Phe Glu Ile Arg Thr Pro Leu Lys Lys
                  115 120 125
          Ile Thr Thr Asp Thr Ile Ala Lys Ile Glu Thr Val Asn Ser Val Ser
              130 135 140
          Ile His Ile Arg Arg Gly Asp Tyr Leu Asn Asn Pro Leu His Asn Thr
          145 150 155 160
          Ser Lys Glu Glu Tyr Tyr Lys Lys Ala Leu Glu Ile Ile Glu Glu Lys
                          165 170 175
          Ile Thr Asn Pro Val Phe Phe Val Phe Ser Asp Asp Ile Asn Trp Val
                      180 185 190
          Lys Ser Asn Phe Ser Thr Asn His Glu Thr Val Phe Ile Asp Phe Asn
                  195 200 205
          Asp Ala Ser Thr Asn Phe Glu Asp Leu Lys Leu Met Ala Ser Cys Lys
              210 215 220
          His Asn Val Ile Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala Trp Leu
          225 230 235 240
          Asn Lys Asn Gln Asn Lys Ile Val Ile Ala Pro Lys Leu Trp Phe Asn
                          245 250 255
          Asp Asn Ser Ile Asn Ile Asn Asp Ile Ile Pro Thr Ser Trp Leu Lys
                      260 265 270
          Ile
           <![CDATA[ <210> 49]]>
           <![CDATA[ <211> 295]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Bacteroidetes]]>
           <![CDATA[ <400> 49]]>
          Met Ile Ile Val Asn Leu Tyr Gly Gly Leu Gly Asn Gln Leu Phe Gln
          1 5 10 15
          Tyr Ala Cys Gly Lys Ala Val Ala Glu Arg Leu Gly Val Glu Leu Gln
                      20 25 30
          Leu Asp Ile Ser His Leu Lys Glu Tyr Gly Lys Ala Thr Asn Phe Thr
                  35 40 45
          Val Arg Asp Phe Glu Leu Gly Val Phe Ser Ile Asn Glu Lys Thr Ala
              50 55 60
          Asp Ile Gln Glu Val Arg Lys Tyr Val Pro Asn Leu Tyr Lys Thr Ser
          65 70 75 80
          Arg Tyr Tyr His Gln Leu Trp Arg Ile Lys Arg Leu Phe Asn Gly Arg
                          85 90 95
          His Leu Phe Ile Glu Arg Thr Trp Gln Arg Asp Arg Tyr Ile Glu Asn
                      100 105 110
          Ile Glu Lys Val Lys Asp Asn Thr Tyr Leu Tyr Gly Tyr Phe Gln Ser
                  115 120 125
          Ala Arg Tyr Phe Ala Asp Ile Glu Glu Lys Leu Arg Thr Thr Leu Val
              130 135 140
          Leu Lys Asp Arg Ile Asp Ala Glu Asn Glu Ala Leu Ile Gln Gln Met
          145 150 155 160
          Glu Asn Glu Asn Ser Val Ser Ile His Ile Arg Arg Gly Asp Tyr Glu
                          165 170 175
          Asn Ser Arg Phe Ser Leu Pro Glu Leu Asp Thr Tyr Tyr Leu Pro Ala
                      180 185 190
          Ile Lys Ala Ile Gln Thr Gln Ile Pro Gln Pro Lys Phe Tyr Ile Phe
                  195 200 205
          Ser Asn Asp Ala Glu Trp Val Lys Lys Asn Phe Asn Ser Leu Glu Ile
              210 215 220
          Lys Asn Glu Ile Val Ser Ile Asn Ser Gly Ser Arg Ser Phe Met Asp
          225 230 235 240
          Met Ile Leu Met Ser Arg Cys Lys His Asn Ile Ile Gly Asn Ser Ser
                          245 250 255
          Phe Ser Trp Trp Gly Ala Trp Leu Asn Gln Asn Pro Asp Lys Met Ile
                      260 265 270
          Val Val Pro Lys Asn Trp Tyr Lys Asn Gly Arg Glu Thr Asp Leu Gln
                  275 280 285
          Leu Lys Glu Trp Ile Lys Ile
              290 295
           <![CDATA[ <210> 50]]>
           <![CDATA[ <211> 279]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Geobacter sp. AJM]]>
           <![CDATA[ <400> 50]]>
          Met Ile Ile Val Lys Leu Gln Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Ala Ala Arg Thr Leu Ser Asn Ser Gly Lys Val Tyr Leu Asp
                      20 25 30
          Phe Ser Phe Leu Asn Lys Asn Asn Ile Thr Thr Asp Ser Phe Thr Ala
                  35 40 45
          Arg Gln Phe Glu Leu Asp Ile Phe Lys Arg Thr Lys Thr His Ile Ser
              50 55 60
          Asn Pro Tyr Phe Ile Arg Leu Ile Arg Ser Asn Arg Lys Ile Phe Lys
          65 70 75 80
          Met Leu Ser Pro Arg Tyr Gln Glu Val Lys Asp Glu Asn Ile Phe Asp
                          85 90 95
          Phe Leu Asn Asn Asn Ser Ser Asn Leu Tyr Leu Asp Gly Tyr Phe Gln
                      100 105 110
          Asn Pro Leu Ile Phe Gln Asp Ile Arg Asn Ile Leu Met Asp Glu Phe
                  115 120 125
          Thr Phe Pro Glu Leu Ser Lys Leu Ser Lys Glu Ile Glu Leu Asn Ile
              130 135 140
          Leu Ser Thr Thr Asn Pro Val Ala Ile His Ile Arg Arg Gly Asp Tyr
          145 150 155 160
          Thr Lys Pro Glu Ile Gln Ser Tyr His Gly Ile Leu Pro Leu Asn Tyr
                          165 170 175
          Tyr Lys Gln Gly Ile Glu Arg Ile Lys Ser Lys Val Glu Asn Pro Ile
                      180 185 190
          Phe Tyr Ile Phe Ser Asp Asp Pro Glu Trp Cys Glu His Asn Leu Ser
                  195 200 205
          Phe Ile Asp Asn Lys His Ile Ile Ala Gly Thr Lys Gln Ala Trp Glu
              210 215 220
          Asp Met Phe Leu Ile Ser Lys Cys Lys His Gln Ile Ile Ala Asn Ser
          225 230 235 240
          Ser Phe Ser Trp Trp Gly Ala Trp Leu Asn Ala Asn Pro Glu Lys Ile
                          245 250 255
          Ile Val Ala Pro Lys Lys Trp Phe Asn Asn Val Asp Ile Asn Ile Leu
                      260 265 270
          Pro Lys Ala Trp Ile Ser Leu
                  275
           <![CDATA[ <210> 51]]>
           <![CDATA[ <211> 317]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Candidate Lakeside Plankton Bacteria]]>
           <![CDATA[ <400> 51]]>
          Met Lys Arg Ser Asn Ser Ser Val Gly Val Ser Leu Thr Gly Gly Leu
          1 5 10 15
          Gly Asn Gln Leu Phe Gln Val Ala Ala Ala Leu Ala Met Ser Glu Gly
                      20 25 30
          Glu Arg Val Thr Leu Ile Ser Ser Tyr Gly Lys Pro Arg Phe Ser Lys
                  35 40 45
          Asn Asn Glu Ala Glu Ile Tyr Ser Leu Ile Arg Ala Glu Asp Trp Phe
              50 55 60
          Glu Arg Asp Asp Glu Val Ser Ser Trp Leu Pro Ala Lys Cys Val Gly
          65 70 75 80
          Tyr Ile Leu Arg Met Gly Val Gln Pro Lys Ile Trp Glu Lys Gly Leu
                          85 90 95
          Val Val Phe Phe Ile Asn Lys Ile Ala Asn Leu Ile Leu Ser Ile Ser
                      100 105 110
          Arg Asn Ser Lys Ile Arg Ile Leu Ala Gly Lys Gly Val Gly Tyr Phe
                  115 120 125
          Thr Leu Pro Glu Lys Lys Tyr Phe Thr Leu Leu Ser Gly Tyr Phe Gln
              130 135 140
          Ser Tyr Arg Trp Ala Ser Leu Asp Asp Val Tyr Lys Lys Leu Phe His
          145 150 155 160
          Leu His Pro Lys His Pro Gly Ser Asp Leu Lys Tyr Tyr Glu Ser Leu
                          165 170 175
          Thr Ser Asp Cys Lys Pro Leu Val Ile His Ile Arg Leu Gly Asp Tyr
                      180 185 190
          Leu Ser Glu Arg Asp Phe Gly Ile Pro Ser Val Glu Tyr Tyr Lys Ser
                  195 200 205
          Ala Val Ser Arg Ile Ser Thr Glu Val Glu Phe Asp Glu Ile Trp Val
              210 215 220
          Phe Ser Asp Thr Leu Thr Lys Ala Lys Glu Ile Leu Arg Phe Asp Gly
          225 230 235 240
          Arg Phe Lys Val Arg Trp Ile Gly Glu Leu Asp Gly Ser Ala Ala Ser
                          245 250 255
          Thr Phe Gln Ala Met Arg His Gly Ala Gly Tyr Ile Ile Gly Asn Ser
                      260 265 270
          Thr Phe Ser Trp Trp Ala Ala Phe Leu Arg Leu Asp Thr Lys Ala Pro
                  275 280 285
          Val Ile Ala Pro Lys Pro Trp Phe Arg Gly Met Glu Glu Pro Phe Glu
              290 295 300
          Leu Ile Pro Lys Asn Trp Thr Arg Ile Asp Ser Asn Tyr
          305 310 315
           <![CDATA[ <210> 52]]>
           <![CDATA[ <211> 304]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Rhizobia bacteria PAR1]]>
           <![CDATA[ <400> 52]]>
          Met Ala Thr Gln Pro Arg Ile Pro Asp Gln Thr Glu Arg Lys Thr
          1 5 10 15
          Gln Val Ala Val Gln Leu His Gly Gly Leu Gly Asn Gln Met Phe Gln
                      20 25 30
          Ala Ala Ala Gly Leu Ala Leu Ala Glu Arg Leu Gly Ala Ser Leu Leu
                  35 40 45
          Phe Asp Thr Ser Arg Phe Arg Asp Lys Gly Leu Arg Ala Tyr Ala Leu
              50 55 60
          Ser Ala Phe Ala Leu Ser Ala Lys Val Leu His Glu Thr Pro Thr Pro
          65 70 75 80
          Leu Ala Ala Leu Arg Arg Ala Ile Leu Lys Gly Met Gly Arg Lys Ser
                          85 90 95
          Ala Thr Gln Pro Asn Tyr Trp Arg Gly Arg Phe Tyr Arg Glu Pro His
                      100 105 110
          Phe His Phe Asp Pro Gly Phe Thr His Leu Ala Gly Asp Thr Met Ile
                  115 120 125
          Ala Gly Tyr Phe Gln Ser Pro His Tyr Phe Ala Gly Tyr Glu Ser His
              130 135 140
          Ile Ala Leu Val Leu Ala Pro Glu Lys Leu Met Ser Asn Ala Ala Leu
          145 150 155 160
          His Leu Ala Glu Ile Leu Asn Gly Glu Ala Ser Val Ala Ile His Leu
                          165 170 175
          Arg Arg Gly Asp Phe Ala Ala Asp Pro Lys Ala Ala Ala Val His Gly
                      180 185 190
          Val Leu Asp Trp Ser Tyr Tyr Asp Arg Ala Val Ala His Ile Arg Ala
                  195 200 205
          Gln Gln Pro Glu Ala Arg Phe Phe Val Phe Ser Asp Asn Pro Glu Ala
              210 215 220
          Ala Ala Glu Gly Ala Ala Arg Trp Pro Asn Ala Glu Pro Met Ala Gly
          225 230 235 240
          Thr Ser Ala Gly Asp Asp Leu Phe Leu Met Ser Arg Ala Arg His His
                          245 250 255
          Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Ser Cys Trp Leu Asp Arg
                      260 265 270
          Arg Glu Gly Gly Ile Arg Ile Ala Pro Glu Gln Trp Phe Ala Ala Gly
                  275 280 285
          Asn Pro Gln Glu Thr Arg Asp Leu Cys Pro Pro Glu Trp Leu Arg Leu
              290 295 300
           <![CDATA[ <210> 53]]>
           <![CDATA[ <211> 396]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Helicobacter sp. 11S02629-2]]>
           <![CDATA[ <400> 53]]>
          Met Ile Glu Ile Arg Leu Gln Gly Arg Leu Gly Asn Gln Phe Phe Ile
          1 5 10 15
          Tyr Ala Phe Ala Lys Asn Leu Arg Lys His Leu Arg Asn Ser Asn Ser
                      20 25 30
          Pro Lys Leu Ala Lys Gln Glu Val Ile Leu Val Cys Asp Asp Ile His
                  35 40 45
          Ser Tyr Glu Val Pro Asp Ile Leu Lys Tyr Asn Val Asp Lys Ser Phe
              50 55 60
          Phe Arg Pro Ser Ala Phe His Gly Phe Lys Thr Thr Ile Asp Lys Ala
          65 70 75 80
          Phe Phe Phe Met Phe Arg Val Ala Arg Lys Gly Ile Tyr Leu Ile Thr
                          85 90 95
          Lys Lys Thr Ile Ser Tyr Arg His Lys Asn Lys Glu Val Pro His Met
                      100 105 110
          Leu Pro Glu Thr Leu Met Glu Thr Tyr Glu Asn Gly Glu Phe Asp Lys
                  115 120 125
          Ala Val Ile Ala Lys Cys Leu Asn Lys Phe Ile Ile Gly Tyr Phe Gln
              130 135 140
          Ser Glu Leu Tyr Phe Glu Asp Val Lys Glu Glu Leu Lys Lys Asp Phe
          145 150 155 160
          Ser Leu Gln Glu Asn His Leu Pro Val Met Lys Val Ser Ala Phe Lys
                          165 170 175
          Ala Leu Leu Thr Lys Ile Lys Glu Gln Asp Ser Ile Phe Leu His Ile
                      180 185 190
          Arg Arg Gly Asp Tyr Leu Ser Ser Val Asn Val Thr Lys Tyr Ala Gln
                  195 200 205
          Leu Gly Val Asn Tyr Tyr Thr Lys Ala Ile Ala Leu Val Leu Gln Lys
              210 215 220
          Cys Pro Asn Ala Lys Phe Tyr Ile Phe Ser Asp Asp Val Ala Trp Val
          225 230 235 240
          Lys Lys Glu Gly Ile Ser Leu Phe Gly Leu Asn Lys Leu Asp Tyr Glu
                          245 250 255
          Val Val Asn Leu Asn Ser Pro Asn Ala Gly Tyr Leu Asp Leu Glu Leu
                      260 265 270
          Met Lys Ala Cys Lys Gly Ala Ile Thr Ala Asn Ser Ser Phe Ser Tyr
                  275 280 285
          Trp Gly Ala Tyr Leu Met Glu Asn Pro Lys Val Val Ile Ala Pro Phe
              290 295 300
          Pro Phe His Phe Ile His Ser Asn Met Asp Leu Leu Pro Arg Asp Phe
          305 310 315 320
          Ile Val Leu Asp Cys Lys Thr Gly Glu Val Leu Glu Glu Lys Asp Tyr
                          325 330 335
          Met Tyr Thr Asn Ser Lys Ser Pro Lys Ser Leu Lys Val Ser Ala Phe
                      340 345 350
          Thr Glu Met Phe Glu Ala Ala Phe Asp Val Val Tyr Asn Pro Asn Ser
                  355 360 365
          Gly Gly Gly Gly Gly Asn Ile Pro Leu Gln Lys Thr Thr Lys Ile Ala
              370 375 380
          Lys Val Val Asn Phe Phe Lys Lys Leu Ile Leu Arg
          385 390 395
           <![CDATA[ <210> 54]]>
           <![CDATA[ <211> 277]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Candidate sire Gassanilacia]]>
           <![CDATA[ <400> 54]]>
          Met Ile Phe Asp Lys Leu Ser Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Ala Gly Tyr Ser Leu Ser Leu Asn Asn Arg Ile Pro Leu Asn
                      20 25 30
          Leu Asp Leu Ser Ser Phe Glu His Lys Lys Thr Gly Ile Thr His Arg
                  35 40 45
          Tyr Phe Leu Leu Asn Lys Phe Asn Ile Asp Arg Asn Ile Leu Ile Asn
              50 55 60
          His Met Glu Lys Ile Ser Gly Tyr Arg Lys Phe Leu Ser Lys Phe Ile
          65 70 75 80
          Thr Lys Phe Phe Gly Glu Asn Phe Tyr Tyr Asn Ile Thr Phe Leu Ser
                          85 90 95
          Ser Lys Tyr Leu Asp Gly Tyr Phe Gln Ser Glu Lys Tyr Phe Lys Asn
                      100 105 110
          Ile Glu Asp Ile Leu Arg Lys Glu Phe Thr Leu Lys Asn Glu Met Ser
                  115 120 125
          Val Val Ala Arg Gln Val Glu Ser Lys Ile Ser Asn Ser Ile Asn Ser
              130 135 140
          Val Ser Leu His Ile Arg Arg Gly Asp Tyr Val Leu Asp Asn Lys Thr
          145 150 155 160
          Asn Ser Tyr His Gly Ile Cys Asp Leu Asp Tyr Tyr Lys Lys Ala Val
                          165 170 175
          Glu Tyr Phe Lys Asn Lys Leu Gly Glu Leu Asn Ile Phe Val Phe Ser
                      180 185 190
          Asp Asp Ile Ala Trp Val Lys Glu Asn Leu Arg Leu Glu Asn Leu Tyr
                  195 200 205
          Phe Val Ser Ser Pro Asp Ile Lys Asp Tyr Glu Glu Leu Ile Leu Met
              210 215 220
          Ser Arg Cys Lys His Asn Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp
          225 230 235 240
          Gly Ala Trp Leu Asn Ala Asn Lys Asn Lys Thr Val Ile Thr Pro Lys
                          245 250 255
          Lys Trp Phe Gln Lys Tyr Asn Ile Asn Gln Lys His Ile Val Pro Lys
                      260 265 270
          Ser Trp Ile Arg Leu
                  275
           <![CDATA[ <210> 55]]>
           <![CDATA[ <211> 284]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Candidate sire Gassanilacia]]>
           <![CDATA[ <400> 55]]>
          Met Ile Ile Val Lys Leu Ser Ala Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Leu Gly Arg Arg Leu Ser Leu Asp Trp Gly Asp Glu Leu Leu
                      20 25 30
          Phe Asp Ser Ser Trp Phe Tyr Ser Thr Arg Glu Gly Glu Thr Pro Arg
                  35 40 45
          Glu Leu Glu Leu His Lys Phe His Leu Leu Leu Pro Glu Ala His Pro
              50 55 60
          Ala Asp Ile Glu Lys Ala Lys Pro Cys Thr Phe Ile Lys Ile Val Lys
          65 70 75 80
          Lys Val Lys Ala Arg Leu Asp Lys Asn Val Phe Tyr Arg Phe Asn Lys
                          85 90 95
          Thr Leu Leu Lys Lys Arg Lys Tyr Val Tyr Leu Asp Gly Tyr Phe Gln
                      100 105 110
          Ser Tyr Lys Tyr Phe Glu Ser Ile Arg Glu Thr Leu Leu Thr Asp Phe
                  115 120 125
          Ile Leu Thr Asp Gly Tyr Ser Asp Asp Ala Tyr Thr Thr Lys Gln Glu
              130 135 140
          Ile Glu Arg Val Gly Glu Ser Val Ser Ile His Ile Arg Arg Gly Asp
          145 150 155 160
          Phe Ala Ser Thr Cys Lys Thr Trp Asn Gly Leu Cys Ser Ile Asp Tyr
                          165 170 175
          Tyr Gln Lys Ala Leu Ala Thr Ile Gln Lys Thr His Pro Leu Val Thr
                      180 185 190
          Val Phe Ile Phe Ser Asp Asp Ile Lys Trp Ala Lys Glu Asn Leu Tyr
                  195 200 205
          Phe Asp Ala Val Pro Met Val Phe Val Ser Arg Pro Ser Leu Ser Ala
              210 215 220
          Thr Glu Glu Leu Ser Leu Met Ser Leu Cys Lys His Gln Ile Ile Ala
          225 230 235 240
          Asn Ser Thr Phe Ser Trp Trp Ala Ala Trp Leu Asn Lys Asn Val Glu
                          245 250 255
          Lys Ile Val Val Ala Pro Ser Arg Trp Leu Val Val Ser Asn Ile Asp
                      260 265 270
          Thr Val Asp Leu Leu Pro Pro Asn Trp Ile Gln Ile
                  275 280
           <![CDATA[ <210> 56]]>
           <![CDATA[ <211> 305]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Aerobic diaminobutyric acid unit bacteria]]>
           <![CDATA[ <400> 56]]>
          Met Asn Lys Ser Val Thr Gln His Thr Glu Ile Asn Ile Gly Asn Gly
          1 5 10 15
          Val Cys Ala Tyr Val Ala Gly Gly Leu Gly Asn Gln Leu Phe Ile Leu
                      20 25 30
          Ala Ala Ala Trp Glu Gln Ala Lys Arg Leu Glu Cys Pro Leu Tyr Leu
                  35 40 45
          Asp Thr Ser Asn His Asp Val Arg Gly Gly Trp Arg Phe Glu Leu Glu
              50 55 60
          Asp Ile Gly Ala Pro Gly Val Val Leu Gly Pro His Ser Pro Trp Thr
          65 70 75 80
          Ser Val Arg Leu Ser Lys Glu Arg Val Leu Pro Phe Pro Arg Phe Thr
                          85 90 95
          Arg Pro Val Arg Arg Arg Val Phe Phe Glu Arg Asp Ser Ser Arg Tyr
                      100 105 110
          Asp Pro Ala Ile Asn Asp Ile Arg Pro Gly Thr Thr Ile Phe Gly Tyr
                  115 120 125
          Phe Gln Ser Ala Arg Tyr Phe Glu Arg Ser Ala Asp Ser Met His Gln
              130 135 140
          Leu Ile Glu Ser Ala Pro Thr Ser Pro Ala Glu Ala Asp Leu Leu Ala
          145 150 155 160
          His Tyr Ala Ala Thr Pro Arg Leu Thr Leu His Leu Arg Arg Gly Asp
                          165 170 175
          Tyr Leu Asn Ala Pro Glu Ser Arg Arg Val Ile Ala Thr Thr Ala Tyr
                      180 185 190
          Ala Arg Arg Ala Tyr Arg Leu Leu Gln Gln Met Gly Ala Ala Ala Pro
                  195 200 205
          Leu Arg Leu Phe Ser Asp Ser Pro Asn Leu Val Gln Glu Glu Leu Gly
              210 215 220
          Asp Leu Asp Leu Asp Ile Glu Phe Ala Asp Pro Glu Gly Val Leu Ser
          225 230 235 240
          Pro Val Asn Thr Met Arg Ala Met Ser Tyr Gly Ser Gly Leu Val Met
                          245 250 255
          Ser Asn Ser Ser Phe Ser Trp Trp Ala Gly Trp Leu Met Arg Arg Arg
                      260 265 270
          Asp Ser Asn Leu Pro Val Ile Ala Pro Arg Pro Trp Asn Glu Thr Gly
                  275 280 285
          Thr Ala Lys Ala Asp Leu Leu Phe Pro Glu Trp Ile Gly Leu Asp Ala
              290 295 300
          Arg
          305
           <![CDATA[ <210> 57]]>
           <![CDATA[ <211> 273]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Flavobacterium megaterium]]>
           <![CDATA[ <400> 57]]>
          Met Ile Val Val Lys Leu Ile Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Ala Ala Lys Ala Ile Ala Ile Glu Arg Asn Gln Pro Leu Glu
                      20 25 30
          Leu Asp Val Ser Ala Phe Asp His Tyr Lys Leu His Gln Phe Ala Leu
                  35 40 45
          His His Phe Asn Phe Ser Ala Lys Val Phe Arg Pro Arg His Ile Leu
              50 55 60
          Trp Asp Lys Val Arg Ser Leu Phe Ile Lys Ile Leu His Tyr Lys Glu
          65 70 75 80
          Arg Asp Phe Gly Phe Asn Ala Arg Ile Phe Asp Leu Lys Ala Asp Asp
                          85 90 95
          Leu Tyr Leu Asp Gly Tyr Phe Gln Ser Glu Lys Tyr Phe Leu Lys His
                      100 105 110
          Ser Ser Thr Ile Arg Arg Glu Phe Glu Ile Ile Ser Ser Leu Lys Pro
                  115 120 125
          Gln Thr Leu Lys Val Leu Glu Glu Ile Arg Asn Thr Asn Ser Val Ser
              130 135 140
          Leu His Ile Arg Arg Gly Asp Tyr Val Gly Asn Pro Leu His Gly Thr
          145 150 155 160
          Asp Asn Glu Leu Phe Tyr Arg Lys Ala Leu Gly Phe Ile Glu Thr Lys
                          165 170 175
          Ile Glu Asn Pro Lys Leu Phe Val Phe Ser Asp Asp Thr Leu Trp Ala
                      180 185 190
          Lys Gln Asn Phe Lys Ser Lys His Glu Val Ile Phe Ile Asp Phe Asn
                  195 200 205
          Asp Ala Leu Ser Asn Phe Glu Asp Ile Lys Leu Met Ser Cys Cys Lys
              210 215 220
          His Asn Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala Trp Leu
          225 230 235 240
          Asn Asp Asn Pro Gly Lys Leu Val Val Ala Pro Gln Gln Trp Phe Asn
                          245 250 255
          Asp Lys Ser Ile Asn Thr Ser Asp Leu Ile Pro Glu Asn Trp Ile Arg
                      260 265 270
          Phe
           <![CDATA[ <210> 58]]>
           <![CDATA[ <211> 283]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Pontilitobacterium]]>
           <![CDATA[ <400> 58]]>
          Met Ile Ile Ala Arg Ile Phe Gly Gly Leu Gly Asn Gln Leu Phe Gln
          1 5 10 15
          Tyr Ala Thr Ala Arg Ala Leu Ala Lys Arg Ala Cys Ala Pro Leu Trp
                      20 25 30
          Leu Asp Ile Arg Leu Ala Pro Pro Gly Asp His Trp Glu Phe Ala Leu
                  35 40 45
          Asn His Phe Ala Ile Glu Ala Lys Ile Ala Gln Pro Gly Glu Leu Pro
              50 55 60
          Pro Asp Lys Asn Ser Lys Leu Arg Tyr Ala Ala Trp Arg Tyr Leu Gly
          65 70 75 80
          Gly Ser Pro Lys Phe Thr Arg Glu Arg Gly Leu Gly Phe Asn Glu Asn
                          85 90 95
          Ile Leu Ser Leu Arg Gly Asp Val Tyr Leu His Gly Tyr Phe Gln Ser
                      100 105 110
          Glu Arg Tyr Phe Glu Asp Phe Pro Gly Leu Arg Gln Glu Leu Thr Ile
                  115 120 125
          Lys Thr Pro Pro Ser Asp Glu Asn Arg Arg Trp Ala Asp Asp Ile Arg
              130 135 140
          Ala Val Pro Ser Val Ser Leu His Leu Arg Arg Gly Asp Tyr Leu Thr
          145 150 155 160
          Ala Lys Gly Ala Gly Ser His Ala Ser Cys Asp Ala Ala Tyr Tyr Glu
                          165 170 175
          Arg Ala Leu Asn His Val Ala Glu Lys Met Asp Ala Glu Pro Lys Val
                      180 185 190
          Phe Val Phe Ser Asp Asp Pro Asp Trp Ala Arg Asp Asn Leu Lys Leu
                  195 200 205
          Pro Phe Glu Met Arg Val Ala Gly His Asn Gly Ser Asp Lys His Tyr
              210 215 220
          Glu Asp Met Arg Leu Ile Ser Ile Cys Arg His Asn Ile Ile Ala Asn
          225 230 235 240
          Ser Thr Phe Ser Trp Trp Gly Ala Trp Leu Asn Asp Asn Pro Asp Lys
                          245 250 255
          Val Val Val Ala Pro Lys Val Trp Phe Ala Asn Pro Lys Leu Ala Asn
                      260 265 270
          Pro Asp Ile Thr Pro Glu Ser Trp Ile Arg Leu
                  275 280
           <![CDATA[ <210> 59]]>
           <![CDATA[ <211> 290]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Kinomonas alginolytica]]>
           <![CDATA[ <400> 59]]>
          Met Ile Thr Leu Ile Ile Ser Gly Gly Leu Gly Asn Gln Met Phe Glu
          1 5 10 15
          Tyr Ala Ala Gly Arg Ala Leu Ser Leu Arg His Arg Thr Asn Leu Ser
                      20 25 30
          Ile Asp Leu Tyr Ile Leu Asn Lys Lys Thr Lys Ala Thr Ile Arg Asn
                  35 40 45
          Tyr Glu Leu Ile Val Phe Asn Ile Glu Thr Pro Ile Thr Ser Ser Ile
              50 55 60
          Phe Asn Lys Ile Ala Val Lys Gly Phe Gly Leu Leu Lys Ser Ser Asn
          65 70 75 80
          Thr Arg Arg Ile Leu Leu Ser Asn Thr Gly Ile Phe Arg Asp Glu Lys
                          85 90 95
          Ala Gln Cys Tyr Asp Ser Arg Phe Lys Lys Leu Ser Glu Lys Met Thr
                      100 105 110
          Leu Phe Gly Tyr Phe Gln Asn Glu Asn Tyr Phe Arg Asp Ile Ser Glu
                  115 120 125
          Gln Leu Arg Thr Asp Phe Thr Phe Gln Ala Pro Leu Ile Gly Arg Asn
              130 135 140
          Asp Glu Ile Arg Ser Leu Ile Glu Leu Asn Thr Ser Val Ser Ile His
          145 150 155 160
          Ile Arg Arg Gly Asp Tyr Ser Asn Thr Asn Ser Asn Leu Pro Ile Leu
                          165 170 175
          Asp Ile Ser His Tyr Lys Lys Ala Ile Glu Tyr Ile Ser Ser Gln Ile
                      180 185 190
          Ser Asn Pro Tyr Phe Phe Ile Phe Ser Asp Asp Ile Asp Trp Val Lys
                  195 200 205
          Asn Asn Leu Asp Leu Ser Asp Ser Asn His Gln Phe Ile Asp Trp Asn
              210 215 220
          Lys Asn Lys Asn Ser Tyr Ile Asp Met Gln Leu Met Ser Leu Cys Lys
          225 230 235 240
          His Asn Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala Trp Leu
                          245 250 255
          Asn Thr Asn Pro Asp Lys Leu Val Ile Ala Pro Asp Lys Trp Tyr Lys
                      260 265 270
          Gly Asp Asn Gly Ile Tyr Pro Asp Gly Phe Leu Pro Lys Glu Trp Ile
                  275 280 285
          Val Leu
              290
           <![CDATA[ <210> 60]]>
           <![CDATA[ <211> 285]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Campylobacter suis subsp. Lawsonia]]>
           <![CDATA[ <400> 60]]>
          Met Val Ile Val Lys Ile Leu Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Tyr Ala Lys Ser Leu Glu Gln Asn Gly Tyr Ile Val Lys Ile
                      20 25 30
          Asp Ile Ser Gly Phe Lys Asp Tyr Val Leu Gln Lys Tyr Gly Leu Asp
                  35 40 45
          Lys Tyr Asn Ile Asp Leu Gln Ile Ser Gln Lys His Glu Asn Asp Lys
              50 55 60
          Ile Tyr Lys Asn Thr Leu Phe Tyr Lys Ile Leu Arg Lys Ile Ser Val
          65 70 75 80
          Asn Leu Ser Ser Ser Ile Lys Glu Lys Asp Leu Arg Phe Asn Asn Lys
                          85 90 95
          Leu Leu Asn Ile Lys Asp Asn Ser Tyr Ile Tyr Gly Tyr Phe Gln Ser
                      100 105 110
          Glu Lys Tyr Phe Lys Asp Ile Arg Glu Ile Ile Leu Lys Gln Phe Thr
                  115 120 125
          Ile Asn Gln Glu Phe Ser Asp Tyr Ala Lys Asp Ile Glu Lys Lys Ile
              130 135 140
          Leu Asn Ser Gln Asn Ser Cys Ser Ile His Ile Arg Arg Gly Asp Tyr
          145 150 155 160
          Val Thr Asn Asn Asn Ile Leu Ile His Gly Met Cys Ser Ile Glu Tyr
                          165 170 175
          Tyr Lys Lys Ser Met Lys Tyr Leu Glu Asn Lys Ile Ala Asn Ile Ser
                      180 185 190
          Tyr Phe Ile Phe Ser Asp Asp Ile Glu Trp Val Lys Asn Asn Leu Pro
                  195 200 205
          Ile Gln Asn Gly Ile Tyr Ile Asp Ser Lys Glu Lys Arg Ile Pro His
              210 215 220
          Glu Asp Ile Tyr Leu Met Ser Leu Cys Asn His Asn Ile Ile Ala Asn
          225 230 235 240
          Ser Thr Phe Ser Trp Trp Gly Gly Trp Leu Asn Gln Asn Lys Asn Lys
                          245 250 255
          Ile Val Ile Ala Pro Lys Ile Trp Phe Ala Asp Lys Lys Leu Gln Glu
                      260 265 270
          Gln Ser Lys Asp Ile Val Cys Gln Glu Trp Ile Lys Ile
                  275 280 285
           <![CDATA[ <210> 61]]>
           <![CDATA[ <211> 290]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Plankton bacteria]]>
           <![CDATA[ <400> 61]]>
          Met Ile Ile Val Glu Leu Lys Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Phe Ala Arg Asn Leu Ala His Ile His Asn Thr Thr Leu Lys
                      20 25 30
          Thr Asp Ile Ser Gly Tyr Ala Cys Ser Ser Gly Met Pro Tyr Ala Leu
                  35 40 45
          Ser Pro Phe Asn Val Gln Glu Asn Phe Ala Thr Pro Asp Glu Ile Gln
              50 55 60
          Ser Phe Thr Thr Pro Lys Gln Thr Ala Ala Gly Lys Trp Ile His Ser
          65 70 75 80
          Leu Leu His Asn His Pro Lys Lys Ala Lys Ser Tyr Ile Arg Val Lys
                          85 90 95
          Tyr Pro His Phe Asn Pro Lys Tyr Leu Lys Leu Pro Asp Asn Val Tyr
                      100 105 110
          Leu Ser Gly Tyr Phe Gln Ser Glu Lys Tyr Phe Ile Asn Ile Ala Asp
                  115 120 125
          Ile Ile Arg Asn Glu Phe Lys Ile Lys Thr Glu Met Asp Ser Arg Asn
              130 135 140
          Gly Asn Ile Ala Glu Thr Ile Gln Asn Ser Gln Ser Val Ser Ile His
          145 150 155 160
          Ile Arg Arg Gly Asp Tyr Ile Thr Asn Pro Lys Ala Ile Gln Thr His
                          165 170 175
          Gly Val Cys Gly Leu Asp Tyr Tyr Asn Arg Cys Val Glu Gln Ile Ser
                      180 185 190
          Leu Lys Ile Lys Ser Pro Arg Phe Phe Val Phe Ser Asp Asp Phe Asp
                  195 200 205
          Trp Ser Arg Lys Asn Ile Lys Leu Pro Asn Asp Ala Ile Tyr Ile Gly
              210 215 220
          His Asn Gly Pro Asn Glu Ala Tyr Lys Asp Leu Gln Leu Met Ser Phe
          225 230 235 240
          Cys Lys Tyr His Ile Thr Ala Asn Ser Thr Phe Ser Trp Trp Gly Ala
                          245 250 255
          Trp Leu Ala Ala Tyr Lys Asp Lys Ile Val Phe Val Pro Lys Lys Trp
                      260 265 270
          Phe Ala Lys Asn Met Asn Thr Glu Gly Met Met Pro Asp Asn Trp Ile
                  275 280 285
          Lys Val
              290
           <![CDATA[ <210> 62]]>
           <![CDATA[ <211> 261]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Akkermansia sp.]]>
           <![CDATA[ <400> 62]]>
          Met Asn Met Glu Arg Lys Thr Gly Leu Met Asn Lys Lys Tyr Val Ser
          1 5 10 15
          Pro Cys Phe Leu Pro Gly Met Arg Leu Gly Asn Ile Met Phe Thr Leu
                      20 25 30
          Ala Ala Ala Cys Ala His Ala Arg Thr Val Gly Val Glu Cys Arg Val
                  35 40 45
          Pro Trp Ala Tyr Asn Asp Ala Ser Leu Met Leu Arg Ser Arg Leu Gly
              50 55 60
          Gly Trp Val Leu Pro Ser Thr Pro Cys Gly Thr Asn Glu Pro Ser
          65 70 75 80
          Trp Gln Glu Pro Ser Phe Ala Tyr Cys Pro Val Pro Ser Arg Ile Arg
                          85 90 95
          Thr Gly Gly Leu Arg Gly Tyr Phe Gln Ser Ala Arg Tyr Phe Gly Gly
                      100 105 110
          Gln Glu Ala Phe Ile Arg Ala Leu Phe Ala Pro Leu Thr Ala Glu Lys
                  115 120 125
          Glu Pro Gly Ala Val Gly Ile His Ile Arg Leu Gly Asp Tyr Arg Arg
              130 135 140
          Leu Arg Asp Lys His Arg Ile Leu Asp Pro Gly Phe Leu Arg Arg Ala
          145 150 155 160
          Ala Gly His Leu Ser Ser Gly Lys Asn Arg Leu Val Leu Phe Ser Asp
                          165 170 175
          Glu Pro Asp Glu Ala Ala Glu Met Leu Ala Arg Val Pro Ala Phe Gly
                      180 185 190
          Arg Phe Ala Leu Glu Ile Asp Arg Gly Ala Pro Cys Glu Ser Leu Arg
                  195 200 205
          Arg Met Thr Ala Met Glu Glu Leu Val Met Ser Cys Ser Ser Phe Ser
              210 215 220
          Trp Trp Gly Ala Trp Leu Gly Asn Thr Arg Lys Val Ile Val Pro Arg
          225 230 235 240
          Asp Trp Phe Val Gly Gly Val Glu Asp Tyr Arg Asp Ile Tyr Leu Pro
                          245 250 255
          His Trp Val Thr Leu
                      260
           <![CDATA[ <210> 63]]>
           <![CDATA[ <211> 304]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Helicobacter sp. MIT 17-337]]>
           <![CDATA[ <400> 63]]>
          Met Phe Gly Tyr Ala Phe Ala Lys Ala Leu Glu Ile Gln Tyr Asn Thr
          1 5 10 15
          Glu Val Lys Leu Asp Ile Leu Phe Tyr Thr Lys Lys Ile Asn Arg Lys
                      20 25 30
          Asn Asn Asn Ile Arg Asn Phe Glu Leu Ser His Phe Asp Val His Ile
                  35 40 45
          Lys Tyr Ala Phe Gly Val Ala Asn Arg Leu Asp Lys Ile Ile Thr Ser
              50 55 60
          Leu Ile Pro Lys Arg Phe Arg Ala Asp Phe Thr Pro Ser Pro Phe Asn
          65 70 75 80
          Leu Ile Lys Asp Asn Lys Asp Leu His Ile Lys Ile Lys Glu Arg Tyr
                          85 90 95
          Pro Phe Pro Lys Asp Thr Tyr Phe Glu Gly Tyr Phe Gln Asn Leu Val
                      100 105 110
          Tyr Phe Asp His Ile Arg Asn Ile Leu Leu His Asp Phe Cys Leu Lys
                  115 120 125
          Thr Pro Leu Asp Leu Lys Asn Lys Gln Leu Gln Asp Tyr Ile Leu His
              130 135 140
          Thr His Asn Ser Val Phe Leu His Ile Arg Arg Gly Asp Tyr Leu Glu
          145 150 155 160
          Tyr Glu His Ser Gly Phe Ile Asn Leu Thr Tyr Thr Asn Tyr Tyr Asn
                          165 170 175
          Thr Ala Leu Lys Thr Ile Gln Glu Lys Leu Gly Lys Ala His Ile Phe
                      180 185 190
          Ile Phe Ser Asn Asp Met Val Trp Cys Lys Glu His Phe Leu Asn Gly
                  195 200 205
          Leu Asp Ser Lys Ile Leu Gln Asp Leu Thr Phe Gln Phe Ala Asp Asn
              210 215 220
          Asn Asn Glu Gly Ser Ala Ala Phe Glu Met Glu Leu Met Arg Ser Cys
          225 230 235 240
          Lys His Gly Ile Ile Ala Asn Ser Thr Phe Ser Trp Trp Ala Ala Tyr
                          245 250 255
          Leu Ile Gln Tyr Pro Asn Lys Ile Ile Val Ala Pro Asn Glu Phe Phe
                      260 265 270
          Thr Ile Val Gln Asp Ser Tyr Ser Val Phe His Asp Tyr Glu Asp Lys
                  275 280 285
          Ile Leu Pro Lys Asn Trp His Arg Ile Pro Leu Thr Lys Glu Asn Lys
              290 295 300
           <![CDATA[ <210> 64]]>
           <![CDATA[ <211> 297]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Geobacter sp. G11]]>
           <![CDATA[ <400> 64]]>
          Met Val Ile Val Lys Leu Met Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Ala Ala Arg Ser Leu Ser Phe Lys Arg Lys Val Tyr Leu Asp
                      20 25 30
          Lys Ser Phe Leu Asp Glu His Lys Val Ser Ser Thr Asp Phe Thr Ala
                  35 40 45
          Arg Asp Tyr Glu Leu Asp Lys Phe Lys Asn Ile Gln Ala Arg Asn Gly
              50 55 60
          Ile Lys Asn Cys Ile Thr Phe Phe Thr Asp Phe Arg Phe Pro Phe Asn
          65 70 75 80
          Ile Leu Arg Tyr Phe Phe Ala Arg Tyr Ala Ile Leu Ile Lys Gln Val
                          85 90 95
          Glu Asn Glu Phe Ile Asp Leu Ala Ser Cys Lys Asn Tyr Thr Leu Ile
                      100 105 110
          Tyr Leu Asp Gly Tyr Phe Gln Ser Glu Lys Tyr Phe Ser His Ile Arg
                  115 120 125
          Lys Gln Leu Leu Lys Glu Phe Glu Phe Pro Gln Leu Asp Thr Glu Asn
              130 135 140
          Thr Ala Ile Arg Asn Lys Ile Ile Arg Ser Glu Asn Thr Val Ser Leu
          145 150 155 160
          His Ile Arg Arg Gly Asp Tyr Asn Ser Ser Glu Ile Ile His Asn Ile
                          165 170 175
          His Gly Val Leu Pro Leu Thr Tyr Tyr His Gln Ala Ile Thr Ala Leu
                      180 185 190
          Thr Leu Lys Tyr Gly Lys Leu Ser Ile Tyr Ile Phe Ser Asp Asp Met
                  195 200 205
          Lys Trp Ala Lys Glu His Leu Leu Ile Asn Asp Gln Glu Val His Tyr
              210 215 220
          Val Asp Leu Asn His Lys Lys Lys His Trp Gln Asp Met Ala Leu Met
          225 230 235 240
          Ser Ala Cys Lys His His His Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp
                          245 250 255
          Gly Ala Trp Leu Ala Asp Thr Gly Gly Asp Thr Phe Ala Pro Leu Lys
                      260 265 270
          Trp Phe Asn Ser Asp Ala Thr Lys Phe Glu Ile Asn Asp Phe Ile Pro
                  275 280 285
          Gln Asn Trp Thr Ile Val Asn Tyr Gly
              290 295
           <![CDATA[ <210> 65]]>
           <![CDATA[ <211> 285]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Brevibacterium brevis]]>
           <![CDATA[ <400> 65]]>
          Met Ile Ala Val Glu Ile Ile Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Thr Ala Lys Ala Met Ala Leu His Arg Arg Asp Asp Leu Leu
                      20 25 30
          Ile Asp Lys Arg Pro Phe Glu Ser Tyr Ser Leu His Ser Phe Ser Leu
                  35 40 45
          Pro His Phe Asn Val Thr Ala Pro Tyr Leu Lys Glu Asp Ile Val Leu
              50 55 60
          Asp Leu Lys Ser Thr Ser Asp Lys Leu Lys Ala Phe Ile Lys Gly Gln
          65 70 75 80
          Lys Asn Tyr Thr Leu Tyr Gln Glu Asn Gly Leu Gly Tyr Asp Ala Ser
                          85 90 95
          Leu Phe Asp Leu Gln Ala Lys Asn Val Tyr Leu Lys Gly Tyr Phe Gln
                      100 105 110
          Ser Glu Lys Tyr Phe Ile Lys Tyr Glu Asp Glu Ile Arg Asn Asp Phe
                  115 120 125
          Glu Ile Val Thr Pro Leu Gln Glu Lys Thr Val Glu Met Leu Thr Ile
              130 135 140
          Ile Asn Glu Val Asn Ala Val Ser Leu His Ile Arg Arg Gly Asp Tyr
          145 150 155 160
          Val Thr Asn Pro Glu Ala Asn Ser Val His Gly Thr Cys Asn Leu Asp
                          165 170 175
          Tyr Tyr Gln Arg Ala Ile Ser His Ile Arg Glu Val Val Glu Asn Pro
                      180 185 190
          Ile Phe Phe Ile Phe Ser Asp Asp Ile Gln Trp Ala Lys Glu Asn Leu
                  195 200 205
          Thr Ile Pro Glu Thr Thr His Phe Ile Asp Phe Thr Asp Ala Ser Thr
              210 215 220
          Asn Tyr Glu Asp Ile Lys Leu Met Ser Thr Cys Lys His Asn Ile Ile
          225 230 235 240
          Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala Trp Leu Asn Lys Asn Lys
                          245 250 255
          Ser Lys Ile Val Ile Ala Pro Ser Lys Trp Phe Asn Val Asp Tyr His
                      260 265 270
          Asn Ala Asp Asp Ile Ile Pro Glu Ser Trp Met Lys Ile
                  275 280 285
           <![CDATA[ <210> 66]]>
           <![CDATA[ <211> 261]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Akkermansia muciniphila]]>
           <![CDATA[ <400> 66]]>
          Met Asn Met Glu Arg Lys Thr Gly Leu Met Asn Lys Lys Tyr Val Ser
          1 5 10 15
          Pro Cys Phe Leu Pro Gly Met Arg Leu Gly Asn Ile Met Phe Thr Leu
                      20 25 30
          Ala Ala Ala Cys Ala His Ala Arg Thr Val Gly Val Glu Cys Arg Val
                  35 40 45
          Pro Trp Ala Tyr Asn Asp Ala Ser Leu Met Leu Arg Ser Arg Leu Gly
              50 55 60
          Gly Trp Val Leu Pro Ser Thr Pro Cys Gly Thr Asn Glu Pro Ser
          65 70 75 80
          Trp Gln Glu Pro Ser Phe Ala Tyr Cys Pro Val Pro Phe Arg Ile Arg
                          85 90 95
          Thr Gly Gly Leu Arg Gly Tyr Phe Gln Ser Ala Arg Tyr Phe Glu Gly
                      100 105 110
          Gln Glu Ala Phe Ile Arg Ala Leu Phe Ala Pro Leu Thr Ala Glu Lys
                  115 120 125
          Glu Pro Gly Ala Val Gly Ile His Ile Arg Leu Gly Asp Tyr Arg Leu
              130 135 140
          Leu Arg Asp Lys His Arg Ile Leu Asp Pro Gly Phe Leu Arg Arg Ala
          145 150 155 160
          Ala Gly His Leu Ser Ser Gly Lys Asn Arg Leu Val Leu Phe Ser Asp
                          165 170 175
          Glu Pro Asp Glu Ala Ala Glu Met Leu Ala Cys Val Pro Ala Phe Gly
                      180 185 190
          Arg Phe Ala Leu Glu Ile Asp Arg Gly Ala Pro Cys Glu Ser Leu Arg
                  195 200 205
          Arg Met Thr Ala Met Glu Glu Leu Val Met Ser Cys Ser Ser Phe Ser
              210 215 220
          Trp Trp Gly Ala Trp Leu Gly Asn Thr Arg Lys Val Ile Val Pro Arg
          225 230 235 240
          Asp Trp Phe Val Gly Gly Val Glu Asp Tyr Arg Asp Ile Tyr Leu Pro
                          245 250 255
          His Trp Val Thr Leu
                      260
           <![CDATA[ <210> 67]]>
           <![CDATA[ <211> 261]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Akkermansia muciniphila]]>
           <![CDATA[ <400> 67]]>
          Met Asn Met Glu Arg Lys Thr Gly Leu Met Asn Lys Lys Tyr Val Ser
          1 5 10 15
          Ser Cys Phe Leu Pro Gly Met Arg Leu Gly Asn Ile Met Phe Thr Leu
                      20 25 30
          Ala Ala Ala Cys Ala His Ala Arg Thr Val Gly Val Glu Cys Arg Val
                  35 40 45
          Pro Trp Ala Tyr Asn Asp Ala Ser Leu Met Leu Arg Ser Arg Leu Gly
              50 55 60
          Gly Trp Val Leu Pro Ser Thr Pro Cys Gly Thr Asn Glu Pro Ser
          65 70 75 80
          Trp Gln Glu Pro Ser Phe Ser Tyr Cys Pro Val Pro Ser Arg Ile Arg
                          85 90 95
          Thr Gly Gly Leu Arg Gly Tyr Phe Gln Ser Ala Arg Tyr Phe Glu Gly
                      100 105 110
          Gln Glu Ala Phe Ile Arg Ala Leu Phe Ala Pro Leu Thr Ala Glu Lys
                  115 120 125
          Glu Pro Gly Ala Val Gly Ile His Ile Arg Leu Gly Asp Tyr Arg Arg
              130 135 140
          Leu Arg Asp Lys His Arg Ile Leu Asp Pro Gly Phe Leu Arg Arg Ala
          145 150 155 160
          Ala Gly His Leu Ser Ser Gly Lys Asn Arg Leu Val Leu Phe Ser Asp
                          165 170 175
          Glu Pro Asp Glu Ala Ala Glu Met Leu Ala Arg Val Pro Ala Phe Gly
                      180 185 190
          Arg Phe Ala Leu Glu Ile Asp Arg Gly Ala Pro Cys Glu Ser Leu Arg
                  195 200 205
          Arg Met Thr Ala Met Glu Glu Leu Val Met Ser Cys Ser Ser Phe Ser
              210 215 220
          Trp Trp Gly Ala Trp Leu Gly Asn Thr Arg Lys Val Ile Val Pro His
          225 230 235 240
          Asp Trp Phe Val Gly Gly Val Glu Asp Tyr Arg Asp Ile Tyr Leu Pro
                          245 250 255
          His Trp Val Thr Leu
                      260
           <![CDATA[ <210> 68]]>
           <![CDATA[ <211> 261]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Akkermansia muciniphila]]>
           <![CDATA[ <400> 68]]>
          Met Asn Met Glu Arg Lys Thr Gly Leu Met Asn Lys Lys Tyr Val Ser
          1 5 10 15
          Pro Cys Phe Leu Pro Gly Met Arg Leu Gly Asn Ile Met Phe Thr Leu
                      20 25 30
          Ala Ala Ala Cys Ala His Ala Arg Thr Val Gly Val Glu Cys Arg Val
                  35 40 45
          Ser Trp Ala Tyr Asn Asp Ala Ser Leu Met Leu Arg Ser Arg Leu Gly
              50 55 60
          Gly Trp Val Leu Pro Ser Thr Pro Cys Gly Thr Asn Glu Pro Ser
          65 70 75 80
          Trp Gln Glu Pro Ser Phe Ser Tyr Cys Pro Val Pro Ser Arg Ile Arg
                          85 90 95
          Thr Gly Gly Leu Arg Gly Tyr Phe Gln Ser Ala Arg Tyr Phe Glu Gly
                      100 105 110
          Gln Glu Ala Phe Ile Arg Ala Leu Phe Ala Pro Leu Thr Ala Glu Lys
                  115 120 125
          Glu Pro Gly Ala Val Gly Ile His Ile Arg Leu Gly Asp Tyr Arg Arg
              130 135 140
          Leu Arg Asp Lys His Arg Ile Leu Asp Pro Gly Phe Leu Arg Arg Ala
          145 150 155 160
          Ala Gly His Leu Ser Ser Gly Lys Asn Arg Leu Val Leu Phe Ser Asp
                          165 170 175
          Glu Pro Asp Glu Ala Ala Glu Met Leu Ala Arg Val Pro Ala Phe Gly
                      180 185 190
          Arg Phe Ala Leu Glu Ile Asp Arg Gly Ala Pro Cys Glu Ser Leu Arg
                  195 200 205
          Arg Met Thr Ala Met Glu Glu Leu Val Met Ser Cys Ser Ser Phe Ser
              210 215 220
          Trp Trp Gly Ala Trp Leu Gly Asn Thr Arg Lys Val Ile Val Pro Arg
          225 230 235 240
          Asp Trp Phe Val Gly Gly Val Glu Asp Tyr Arg Asp Ile Tyr Leu Pro
                          245 250 255
          His Trp Val Thr Leu
                      260
           <![CDATA[ <210> 69]]>
           <![CDATA[ <211> 319]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Chitinophaga bacteria]]>
           <![CDATA[ <400> 69]]>
          Met Glu Lys Leu Phe Arg Ile Gln Ala Ala His Arg Leu Ser Gly Val
          1 5 10 15
          Arg Ala His Leu Ile Arg Arg His Leu Phe Gln Lys Leu Ile Ser Arg
                      20 25 30
          Arg Asp Ser Met Ile Ile Val Gln Leu Lys Gly Gly Leu Gly Asn Gln
                  35 40 45
          Met Phe Gln Tyr Ala Ala Ala Arg Ala Leu Ala Ala Arg His Arg Thr
              50 55 60
          Gln Val Leu Leu Asp Thr Ala His Tyr Arg Ala Asp Gln Leu Arg Asn
          65 70 75 80
          Phe Asp Leu Phe His Leu Gln Val Ala Ala Ser Thr Ala Ala Pro Glu
                          85 90 95
          Asp Cys Ala Pro Leu Lys Ala Gln Ser Thr Phe Ser Arg Leu Ala Ala
                      100 105 110
          Arg Leu Thr Pro Tyr Ser Arg Lys Arg Phe Tyr Lys Gln Pro Phe Tyr
                  115 120 125
          His Phe Asp Pro His Phe Phe Glu Leu Gly Pro Asp Val Tyr Leu Gln
              130 135 140
          Gly Tyr Phe Gln Ser Glu Arg Tyr Phe Ala Pro Leu Ala Pro Gln Leu
          145 150 155 160
          Arg Glu Glu Phe Arg Phe Arg Asp Ala Phe Pro Glu His Val Arg Asn
                          165 170 175
          Ala Ala Gly Glu Met Arg Ala Gly Pro Ser Val Ser Leu His Ile Arg
                      180 185 190
          Arg Gly Asp Tyr Arg Asn Pro Glu Thr Leu Arg Val His Gly Ile Leu
                  195 200 205
          Pro Phe Ala Tyr Tyr Glu Ala Ala Ile Arg Lys Leu Gln Glu Arg Ile
              210 215 220
          Gly Pro Pro Ala Phe Tyr Val Phe Thr Asp Asp Pro Gln Trp Val Ala
          225 230 235 240
          Ala Asn Leu Asp Ile Pro Gly Ala Arg Ile Val Ser Gly Ala Thr Ser
                          245 250 255
          Ser Thr His Phe Glu Asp Leu Tyr Leu Met Ser His Cys Arg His His
                      260 265 270
          Ile Leu Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala Trp Leu Asn Thr
                  275 280 285
          Asp Pro Gly Lys Ile Val Ile Ala Pro Arg Gln Trp Phe Asn Glu Gly
              290 295 300
          Pro Val Asp Leu Gln Asp Leu Met Pro Ala Gly Trp Glu Arg Ile
          305 310 315
           <![CDATA[ <210> 70]]>
           <![CDATA[ <211> 284]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Chitinophaga bacteria]]>
           <![CDATA[ <400> 70]]>
          Met Ile Ala Ile Gln Leu Lys Gly Gly Leu Gly Asn Gln Leu Phe Gln
          1 5 10 15
          Tyr Ala Ala Ala Arg Thr Leu Ala Ile Arg Gln His Thr Glu Val Leu
                      20 25 30
          Leu Asp Thr Ser His Tyr Arg Glu Asp Gln Leu Arg Asp Phe Asp Leu
                  35 40 45
          Phe His Leu Asn Val Lys Ala Arg Ile Ala Thr Ala Ser Gln Ile Asp
              50 55 60
          Glu Ile Arg Pro Thr Gly Thr Val Gly Arg Ala Leu Ser His Ile Ala
          65 70 75 80
          Pro Tyr Arg Phe Lys Arg Ile Tyr Gln Gln Pro Phe Tyr His Tyr Asp
                          85 90 95
          Ala Gly Phe Val Arg Leu Arg Gly Pro Val Tyr Leu Lys Gly Tyr Phe
                      100 105 110
          Gln Ser Glu Ala Tyr Phe Arg Pro Val Ala Ala Glu Ile Arg Arg Glu
                  115 120 125
          Leu Thr Phe Arg Glu Leu Pro Leu Pro Gln Ile Gln Ala Leu Gly Ala
              130 135 140
          Arg Leu Arg Gln Glu Gln Ser Val Ser Leu His Ile Arg Arg Gly Asp
          145 150 155 160
          Tyr Lys Asn Pro Glu Thr Leu Arg Val His Gly Ile Leu Pro Leu Glu
                          165 170 175
          Tyr Tyr Arg Ala Ala Val Lys Arg Ile Thr Asp Ser Arg Pro Asp Ala
                      180 185 190
          Arg Phe Tyr Val Phe Thr Asp Asp Leu Glu Trp Val Arg Gln His Leu
                  195 200 205
          Ala Met Pro Glu Ala Met Leu Val Ser Gly Glu Ile Thr Lys Thr His
              210 215 220
          Phe Glu Asp Leu Tyr Leu Met Gln His Cys Arg Asn His Ile Leu Ala
          225 230 235 240
          Asn Ser Ser Phe Ser Trp Trp Gly Ala Trp Leu Asn Pro Asp Lys Glu
                          245 250 255
          Lys Ile Val Val Ala Pro Lys Ala Trp Phe Asn Glu Gly Pro Lys Asp
                      260 265 270
          Thr Gln Asp Leu Ile Pro Arg Ser Trp Ile Arg Leu
                  275 280
           <![CDATA[ <210> 71]]>
           <![CDATA[ <211> 245]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Proteobacteria]]>
           <![CDATA[ <400> 71]]>
          Met Asp Lys Phe Ser Val Leu Thr Ser Gln Ala Ser Pro Ala Glu Val
          1 5 10 15
          Glu Lys Arg Phe Pro Leu Leu Ser Phe Arg Ser Leu Ile Lys Ile Tyr
                      20 25 30
          Arg Arg Leu Leu Arg Ile Leu Pro Lys Leu Ser Arg Lys Tyr Tyr Phe
                  35 40 45
          Glu Lys Ser Met Ala Phe Asp His Ser Leu Phe Glu Thr Asn Pro Glu
              50 55 60
          Cys Phe Ile Asp Gly Tyr Phe Gln Ser Tyr Lys Tyr Phe Glu Gly Val
          65 70 75 80
          Asn Asp Ile Ile Leu Gln Glu Phe Gln Pro Lys Glu Ala Pro Asp Glu
                          85 90 95
          Val Asn His His Leu Ile Arg Val Met Gln Val Ala Asn Ser Val Ser
                      100 105 110
          Leu His Ile Arg Arg Gly Asp Tyr Ile Ser Ser Ala Glu Ala Asn Lys
                  115 120 125
          Val His Gly Ile Cys Gly Leu Glu Tyr Tyr Arg Ala Ala Val Glu Asn
              130 135 140
          Leu Gln Ser Arg Leu Val Asp Pro Gln Tyr Phe Val Phe Ser Asp Asp
          145 150 155 160
          Ile Ala Trp Ala Lys Glu Asn Leu Asp Leu Gly Ala Gly Ala Thr Phe
                          165 170 175
          Val Gln His Asn Ala Gly Ser Lys Ser Tyr Trp Asp Leu Val Leu Met
                      180 185 190
          Ser His Cys Lys His Asn Ile Ile Ala Asn Ser Thr Phe Ser Trp Trp
                  195 200 205
          Gly Ala Trp Leu Asn Gln Asn Pro Asp Lys Ile Val Tyr Ala Pro Lys
              210 215 220
          Thr Trp Phe Ala Asp Ser Ser Lys Asn Thr Ser Asp Leu Ile Pro Gln
          225 230 235 240
          Ser Trp Met Arg Ile
                          245
           <![CDATA[ <210> 72]]>
           <![CDATA[ <211> 339]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Flavobacterium estuarineus sp. 17J68-12]]>
           <![CDATA[ <400> 72]]>
          Met Ser Trp His Leu Arg Val Val Leu Thr Ile Ala Ser Ala Asn Gly
          1 5 10 15
          Cys Asn Gly Arg Cys Arg Asn Gly Cys Pro Thr Arg Ser Leu Cys Cys
                      20 25 30
          Ile Ser Ala Arg Arg Arg Ser Leu Arg His His Ser Glu Thr Phe Ile
                  35 40 45
          Ser Lys Ala Pro Gly Arg Phe Met Ile Val Val Glu Leu Lys Gly Gly
              50 55 60
          Leu Gly Asn Gln Leu Phe Gln Tyr Ala Ala Ala Arg Ala Leu Ala Ala
          65 70 75 80
          Lys His Gly Thr Gly Val Ala Phe Asp Ala Thr His Tyr His Glu Asp
                          85 90 95
          Gln Leu Arg Asn Phe Glu Leu Leu His Met Lys Val Ala Ala Arg Lys
                      100 105 110
          Ala Thr Ser Gly Glu Val Ala Ala Leu Lys Pro His Ser Phe Phe Asp
                  115 120 125
          Arg Leu Arg Asn Arg Leu Ala Pro Tyr Glu Arg Lys Arg Phe Tyr Lys
              130 135 140
          Gln Pro Phe Phe His Tyr Asp Lys Gln Phe Phe Arg Leu Pro Ala Lys
          145 150 155 160
          Val Tyr Leu Arg Gly Tyr Phe Gln Ser Ala Arg Tyr Phe Glu Pro Val
                          165 170 175
          Ala Glu Ile Ile Arg Gln Glu Phe Arg Phe Gln Gln Pro Met Pro Pro
                      180 185 190
          Ala Val Glu Glu Leu Gly Arg Arg Leu Arg Ser Glu Glu Ser Val Ser
                  195 200 205
          Ile His Ile Arg Arg Gly Asp Tyr Lys Asn Pro Glu Thr Leu Arg Val
              210 215 220
          His Gly Ile Gln Pro Leu Ala Tyr Tyr Glu Ala Ala Val Gln Leu Leu
          225 230 235 240
          Arg Glu Arg Ile Asn Ala Pro Leu Phe Tyr Ile Phe Thr Asp Asp Pro
                          245 250 255
          Gly Trp Val Arg Lys Asn Leu Ser Ile Glu Gly Ala Gln Leu Val Ser
                      260 265 270
          Gly Pro Val Ser Ala Thr His Phe Glu Asp Leu Tyr Leu Met Gln Gln
                  275 280 285
          Cys Arg His Asn Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala
              290 295 300
          Trp Leu Asn Asp Asn Ala Gln Lys Met Val Ile Ala Pro Asn Thr Trp
          305 310 315 320
          Phe Asn Glu Gly Pro Lys Asp Thr Gln Asn Leu Ile Pro Gly Thr Trp
                          325 330 335
          Leu Arg Leu
           <![CDATA[ <210> 73]]>
           <![CDATA[ <211> 287]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Geobacter sp. AR-2-6]]>
           <![CDATA[ <400> 73]]>
          Met Ile Val Ile Val Lys Leu Met Gly Gly Leu Gly Asn Gln Met Phe
          1 5 10 15
          Gln Tyr Ala Ala Ala Arg Ser Leu Ser Ser Arg Lys Ile Tyr Phe Asp
                      20 25 30
          Asp Ser Phe Leu Asn Lys Asn Asn Ile Ser Ser Glu Asp Phe Thr Ala
                  35 40 45
          Arg Arg Phe Glu Leu His Ile Phe Arg Arg Leu Lys Val Lys Ile Leu
              50 55 60
          Asn Pro Tyr Ala Gln Arg Leu Leu Leu Thr Lys Asn Lys Lys Tyr Asp
          65 70 75 80
          Phe Leu Lys Leu Leu Leu Pro Asn Ser Leu Lys Arg Ile Cys His Ile
                          85 90 95
          Asp Asp Ser Asn Ile Asn Glu Asn Leu Ser Lys Lys His Gln Asn Glu
                      100 105 110
          Ile Leu Tyr Ile Asp Gly Tyr Phe Gln Asn Pro Thr Tyr Phe Asn Glu
                  115 120 125
          Ile Arg Lys Ser Leu Leu Asn Glu Phe Ala Phe Pro Glu Leu Asp Asp
              130 135 140
          Lys Phe Thr Gly Leu Leu Asn Asp Ile Glu Asn Ser Asn Ser Val Ala
          145 150 155 160
          Ile His Ile Arg Arg Gly Asp Tyr Leu Lys Ser Lys Ile Asn Glu His
                          165 170 175
          His Gly Val Leu Pro Leu Ser Tyr Tyr Lys Asn Ala Val Lys Ile Leu
                      180 185 190
          Glu Gly Lys Leu Ile Asp Pro Lys Tyr Phe Ile Phe Ser Asp Asp Pro
                  195 200 205
          Asn Trp Cys Lys Asn Asn Phe Ala Phe Leu Ala Asn Lys Glu Ile Val
              210 215 220
          Ser Lys Glu Asn Glu Pro Trp Ile Asp Met Tyr Leu Ile Ser Lys Cys
          225 230 235 240
          Lys Asn Gln Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala Trp
                          245 250 255
          Leu Asn Gln Phe Ile Asn Lys Ile Val Ile Ala Pro Lys Asn Trp Phe
                      260 265 270
          Asn Ser Ile Glu Ser Asn Ile Val Pro Lys Glu Trp Ile Ser Leu
                  275 280 285
           <![CDATA[ <210> 74]]>
           <![CDATA[ <211> 246]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Helicobacter marmoset]]>
           <![CDATA[ <400> 74]]>
          Met Tyr Ala Leu Leu Arg Tyr Gly Ile Leu Lys Gly Asn Phe Ala Phe
          1 5 10 15
          Lys Pro Asp Ile Phe Ala Leu Tyr Lys Tyr Arg Tyr Asp Glu His Leu
                      20 25 30
          Ser His Lys Asp Asp Arg Val Trp Gln Ala Phe Thr Gln Ser Ala Lys
                  35 40 45
          Gly Ser Phe His Pro His Ala Leu Pro Ile Gly Tyr Phe Gln Asn Leu
              50 55 60
          Thr Tyr Leu Gln Gly Leu Asp Ser Ile Leu His Lys Glu Phe Cys Leu
          65 70 75 80
          Lys Thr Pro Leu Thr Ala Gln Asn Gln Ala Leu Lys Ala Tyr Ile His
                          85 90 95
          Ser Leu Pro Gln Ser Ala Phe Leu His Ile Arg Leu Gly Asp Tyr Leu
                      100 105 110
          Glu Gly Gly Thr Phe Val Arg Leu Gly Ser Ala Tyr Tyr Asn Gln Ala
                  115 120 125
          Val Gln Ile Leu Lys Lys His Leu Gly Lys Ala Tyr Ile Phe Val Phe
              130 135 140
          Ser Asn Asn Ile Pro Trp Cys Glu Arg Asn Ala His Lys Tyr Ile Asp
          145 150 155 160
          Phe Ser Gly Leu Lys Val Glu Phe Val Lys His Asn Asp Glu Gly Asn
                          165 170 175
          Ala Ala Gln Glu Leu Glu Leu Met Arg Ala Cys Lys His Gly Ile Met
                      180 185 190
          Ala Asn Ser Thr Phe Ser Trp Trp Ala Ala Tyr Leu Asn Asp Asn Pro
                  195 200 205
          His Lys Ile Val Cys Met Pro Lys Tyr Phe Phe Asn Asp Ile Thr Arg
              210 215 220
          Ile Pro Gln Ser Gln Met Ile Ala Lys Gln Asn Tyr Ile Leu Leu Asp
          225 230 235 240
          His Thr Trp Ala Glu Ile
                          245
           <![CDATA[ <210> 75]]>
           <![CDATA[ <211> 295]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Helicobacter japonicum]]>
           <![CDATA[ <400> 75]]>
          Met Gly Gly Leu Gly Asn Gln Met Phe Gln Tyr Ala Phe Ala Lys Ser
          1 5 10 15
          Leu Glu His His Leu Gln Ile Pro Ile Leu Leu Asp Lys Ser Trp Tyr
                      20 25 30
          Asp Asn Pro Lys Asn Ala Lys Met Leu Ser Leu Asp Ile Phe His Met
                  35 40 45
          Asp Leu Val Tyr Ala Thr Lys Thr Gln Val Ile Glu Ala Leu Asn His
              50 55 60
          Pro Asn Ile Glu Gln Leu Thr Glu Ser Arg Ala Phe Glu Arg Lys Pro
          65 70 75 80
          Lys Ile Leu Arg Ser Ile Leu Lys Trp Leu Gly Tyr Lys Lys Pro Thr
                          85 90 95
          Phe Ser Thr Pro Pro Phe Glu Tyr Arg Ala Glu Tyr Leu Lys Ser Asn
                      100 105 110
          Asn Ile Thr Tyr Phe His Gly Tyr Phe Gln Asn Pro Arg Tyr Tyr Gln
                  115 120 125
          Gly Ile Asp Asp Tyr Ile Lys Glu Ser Leu Ser Pro Pro Pro Ile Lys
              130 135 140
          Ser Leu Glu Asn Leu Ser Lys Leu Asp Gln Ile Leu Asn Thr Lys Asn
          145 150 155 160
          Ser Ile Phe Ile His Ile Arg Arg Gly Asp Tyr Leu Ser Leu Ser Trp
                          165 170 175
          Gln Ile Asp Ile Glu Tyr Tyr Lys Lys Ala Val Ala Thr Ile Val Asn
                      180 185 190
          Lys Ile Glu Asn Pro His Phe Phe Leu Phe Cys Ala Asp Arg Glu Phe
                  195 200 205
          Ala His Asn Leu Asp Leu Gly Tyr Pro Phe Ile Asp Met Thr Ser Glu
              210 215 220
          Asn Ile Ser Leu Asp Asn His Phe Glu Asp Leu Ile Leu Met Ala His
          225 230 235 240
          Cys Gln His Gly Ile Val Ala Asn Ser Ser Tyr Ser Trp Trp Ala Ala
                          245 250 255
          Tyr Leu Ile Glu Asn Pro His Lys Ile Ile Ile Ala Pro Thr Pro Trp
                      260 265 270
          Leu His Ser Asn Asp Glu Ile Ile Cys Asp Asp Trp Ile Lys Ile Gln
                  275 280 285
          Ala Glu Lys Glu Val Arg Leu
              290 295
           <![CDATA[ <210> 76]]>
           <![CDATA[ <211> 268]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Klebsiella pneumoniae]]>
           <![CDATA[ <400> 76]]>
          Met Leu Phe Gln Val Cys Leu Ala Met His Leu Arg Asp Met Gly His
          1 5 10 15
          Val Val Lys Ile Asp Thr Leu Gly Leu Glu Asn Lys Phe Lys Asp Lys
                      20 25 30
          Val Met Phe Phe Leu Phe Lys Cys Gly Ile Thr Leu Glu Glu Cys Thr
                  35 40 45
          Lys Ala Glu Arg Phe Ile Cys Cys His Ala Val Ser Thr Asn Arg Ile
              50 55 60
          Lys Ser Lys Glu Leu Lys Ile Ile Lys Leu Ile Phe Pro Lys Lys Leu
          65 70 75 80
          Tyr Leu Glu Gln Gln Trp Gly Glu Ile Pro Asp Asp Ser Tyr Asn Tyr
                          85 90 95
          Tyr Phe Gly Tyr Phe Gln Asn Ile Asn Leu Ala Lys Lys Tyr Cys Asp
                      100 105 110
          Phe Phe Gln Lys Gly Leu Asp Leu Ile Ala Glu Glu Asn Ser Phe Val
                  115 120 125
          Gln Pro Leu Ser Asn Asp Cys Phe Ile His Ile Arg Arg Gly Asp Tyr
              130 135 140
          Cys Thr Pro Ser Ala Leu Ala Leu His Gly Leu Met Gly Glu Glu Tyr
          145 150 155 160
          Tyr Asn Gly Ala Val Arg Tyr Phe Ser Glu Asp Met His Phe Asp Val
                          165 170 175
          Phe Ser Asn Asp Val Ser Trp Val Lys Ser Asn Leu Ser Ile Ser Asn
                      180 185 190
          Ile Lys Val Met Glu Gly Glu Gly Phe Lys Tyr Pro Asp Ile Glu Asp
                  195 200 205
          Leu Tyr Lys Met Ser Arg His Lys Tyr Gly Ile Ile Ala Asn Ser Thr
              210 215 220
          Phe Ser Phe Trp Ala Ala Leu Ile Gly Ser Ser Asn Ile Asn Lys Glu
          225 230 235 240
          Ile Val Cys Pro Glu Lys Trp Phe Ala Asn Glu Glu Leu Gln Lys Leu
                          245 250 255
          Ser Tyr Lys Ile Lys Asn Ser Asp Trp Val Tyr Lys
                      260 265
           <![CDATA[ <210> 77]]>
           <![CDATA[ <211> 293]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Macrophages phagocytosing Abnormalmonas]]>
           <![CDATA[ <400> 77]]>
          Met Ile Arg Val Ile Leu Ser Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Ala Gly Arg Ala Leu Ser Leu Lys Leu Gln Ser Glu Leu Ser
                      20 25 30
          Val Asp Thr Phe Leu Leu Arg Lys Lys Thr Lys Thr Thr Val Arg Asn
                  35 40 45
          Phe Glu Leu Lys Val Phe Asn Ile Lys Val Arg Glu Asn Asn His Leu
              50 55 60
          Lys Asn Lys Leu Ile Thr Lys Ser Phe Phe Phe Leu Asn Lys Tyr Gly
          65 70 75 80
          Leu Lys Lys Leu Ile Phe Gly Leu Phe Gly Ile Phe Arg Asp Gln Lys
                          85 90 95
          Ala Gln Asn Phe Asp Gln Arg Phe Asn Asn Ile Asp Arg Asn Ile Thr
                      100 105 110
          Leu Phe Gly Tyr Phe Gln Asn Glu Asn Tyr Phe Lys Ala Ile Ser Ala
                  115 120 125
          Asn Leu Arg Glu Asp Phe Arg Phe Ile Ser Pro Leu Leu Gly Glu Asn
              130 135 140
          Ala Asp Ile Ala Asn Lys Ile Gly Ser Thr Ala Ser Val Ser Ile His
          145 150 155 160
          Ile Arg Arg Gly Asp Tyr Leu Asn Pro Asn Val Asn Leu Ser Leu Leu
                          165 170 175
          Asn Ile Glu Tyr Tyr Gln Arg Ala Ile Ser Tyr Ile Gln Glu Lys Ile
                      180 185 190
          Thr Asp Pro Val Phe Tyr Ile Phe Ser Asp Asp Ile Asn Trp Val Lys
                  195 200 205
          Lys Asn Leu Asp Leu Ser Gln Ser Pro His Ile Phe Ile Asp Trp Asn
              210 215 220
          Thr Gly Asn Arg Ser Phe Leu Asp Met Gln Leu Met Ser Leu Cys Gln
          225 230 235 240
          His Asn Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala Trp Leu
                          245 250 255
          Asn Cys Asn Pro Asp Lys Ile Val Ile Ala Pro Gly Thr Trp Tyr Lys
                      260 265 270
          Asn Glu Thr Ala Ala Asp Tyr Pro Ile Gly Phe Ile Pro Glu Lys Trp
                  275 280 285
          Val Ile Leu Glu Thr
              290
           <![CDATA[ <210> 78]]>
           <![CDATA[ <211> 204]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Flavobacterium sp.]]>
           <![CDATA[ <400> 78]]>
          Met Gln Asn Asp Asn Glu Ser Cys Glu Met Val Leu Pro Leu Phe Gly
          1 5 10 15
          Asn Trp Lys Phe Tyr Tyr Gly Tyr Phe Gln Ser Glu Gln Tyr Phe Lys
                      20 25 30
          Glu Ile Ala Asp Asn Ile Arg Asp Tyr Phe Arg Ile Lys Ser Ile Tyr
                  35 40 45
          Thr Lys Ala Phe Trp Gln Lys Tyr Gly Tyr Leu Phe Ser Gln Lys Val
              50 55 60
          Leu Ala Ile His Ile Arg Leu Gly Asp Tyr Leu Thr Trp Gly Asn Glu
          65 70 75 80
          Ala Met Gly Gly Glu Asn Met Thr Leu Pro Asp Ala Tyr Phe Gln Asn
                          85 90 95
          Ala Leu Gly Met Ile Pro Asn Ile Gln Asp Tyr Thr Val Leu Val Ile
                      100 105 110
          Thr Asp Asp Ile Glu Asn Ala Ser His Lys Met Gln Trp Leu Gln Gly
                  115 120 125
          Lys Lys Ile Ile Ser Asp Thr Glu Ile Met Asp Phe Gln Leu Leu Met
              130 135 140
          His Ala Asp Lys Leu Ile Ile Ser Asn Ser Ser Phe Ala Trp Trp Ala
          145 150 155 160
          Ala Tyr Leu Asn Val Lys Asn Ala Glu Thr Phe Ala Pro Glu Tyr Trp
                          165 170 175
          Leu Gly Phe Lys Val Asn Thr Glu Ile Pro Asn Asn Val Ile Pro Ser
                      180 185 190
          Arg Phe Lys Arg Val Ala Val Ala Thr Cys Tyr Ala
                  195 200
           <![CDATA[ <210> 79]]>
           <![CDATA[ <211> 291]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Geobacter sp.]]>
           <![CDATA[ <400> 79]]>
          Met Ile Thr Ile Lys Leu Gln Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Ala Ala Arg Ser Leu Ser Lys Lys Ile Phe Leu Asp Leu Asp
                      20 25 30
          Phe Leu Glu Val Asn Cys Thr Asp Lys Leu His Phe Thr Ala Arg Thr
                  35 40 45
          Phe Gln Leu Gly Ile Phe Glu Asn Leu Lys Ile Lys Arg Ser Ser Thr
              50 55 60
          Asn Asn Leu Leu Arg Gln Asn Asn Ile Ala Phe Arg Val Leu Gly Lys
          65 70 75 80
          Leu Phe Lys Ser Arg Ile Ile His Ile Lys Gln Thr Thr Asn Asp Phe
                          85 90 95
          Ile Asn Leu Pro Gly Ser Gly Asp Leu Lys Tyr Ile Tyr Leu Asp Gly
                      100 105 110
          Tyr Phe Gln Ser Glu Lys Tyr Phe Ser His Ile Lys Lys Glu Ile Arg
                  115 120 125
          Asp Glu Phe Lys Phe Pro Ala Leu Asp Ile Lys Asn Leu Asp Ile Ala
              130 135 140
          Asn Gln Ile Arg Asn Ser Glu Asn Ala Thr Ser Ile His Ile Arg Arg
          145 150 155 160
          Gly Asp Tyr Val Lys Ser Ser Ile Val Asn Asp Ile His Gly Thr Leu
                          165 170 175
          Pro Lys Ile Tyr Tyr Gln Lys Ala Val Ser Lys Leu Leu Leu Asp Phe
                      180 185 190
          Pro Lys Ile Ser Ile Phe Val Phe Ser Asp Asp Ile Lys Trp Ala Arg
                  195 200 205
          Lys His Leu Asn Phe Glu Asn Cys Asn Phe Ile Ser Asn Asn Ile Lys
              210 215 220
          Glu Glu Ser Trp Lys Asp Met Ala Leu Met Ser Leu Cys Lys His His
          225 230 235 240
          Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala Trp Leu Ser Ile
                          245 250 255
          Tyr Glu Gly Gln Lys Tyr Ala Pro Val Asn Trp Phe Asn Pro Lys Lys
                      260 265 270
          Ile Lys Tyr Asn Leu Asn Asp Ile Ile Pro Arg Glu Trp Asn Ile Ile
                  275 280 285
          Asp Tyr Asp
              290
           <![CDATA[ <210> 80]]>
           <![CDATA[ <211> 291]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Phototrophic Heflerella]]>
           <![CDATA[ <400> 80]]>
          Met Thr Val Arg Arg Val Ile Ser Arg Met His Gly Gly Leu Gly Asn
          1 5 10 15
          Gln Leu Phe Gln Tyr Ala Val Gly Arg Ala Val Ala Leu Arg Thr Gly
                      20 25 30
          Ser Glu Leu Leu Leu Asp Thr Arg Glu Phe Thr Ser Ser Asn Pro Phe
                  35 40 45
          Gln Tyr Asp Leu Gly His Phe Ser Ile Gln Ala Lys Val Ala Asn Ser
              50 55 60
          Ser Glu Leu Pro Pro Gly Lys Asn Arg Pro Leu Ala Tyr Ala Trp Trp
          65 70 75 80
          Arg Lys Phe Gly Arg Ser Pro Arg Phe Val Arg Glu Gln Asp Leu Gly
                          85 90 95
          Tyr Asn Ala Arg Ile Glu Thr Ile Glu Ala Asp Cys Tyr Leu His Gly
                      100 105 110
          Tyr Phe Gln Ser Gln Lys Tyr Phe Glu Asp Ile Ala Ser Ile Leu Trp
                  115 120 125
          Lys Asp Leu Ser Phe Arg Gln Ala Ile Ser Gly Glu Asn Ala Ser Met
              130 135 140
          Ala Glu Arg Ile Gln Ser Ala Pro Ser Val Ser Met His Ile Arg Arg
          145 150 155 160
          Gly Asp Tyr Leu Thr Ser Ala Lys Ala Arg Ser Thr His Gly Ala Pro
                          165 170 175
          Asp Leu Gly Tyr Tyr Gly Arg Ala Leu Gly Glu Ile Arg Ala Arg Ser
                      180 185 190
          Gly Ser Asp Pro Val Val Tyr Leu Phe Ser Asp Asp Pro Asp Trp Val
                  195 200 205
          Arg Asn Asn Met Arg Met Asp Ala Asn Leu Val Thr Val Ala Ile Asn
              210 215 220
          Asp Gly Lys Thr Ala Phe Glu Asp Leu Arg Leu Met Ser Leu Cys Asp
          225 230 235 240
          His Asn Ile Ile Val Asn Ser Thr Phe Ser Trp Trp Gly Ala Trp Leu
                          245 250 255
          Asn Pro Ser Leu Asp Lys Ile Val Val Ala Pro Lys Arg Trp Phe Ala
                      260 265 270
          Asp Pro Lys Leu Ser Asn Pro Asp Ile Thr Pro Pro Gly Trp Leu Arg
                  275 280 285
          Leu Gly Asp
              290
           <![CDATA[ <210> 81]]>
           <![CDATA[ <211> 305]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Chlorella variably rare]]>
           <![CDATA[ <400> 81]]>
          Met Ile Tyr Ala Glu Leu Ala Gly Gly Leu Gly Asn Gln Met Phe Ile
          1 5 10 15
          Tyr Ala Phe Ala Arg Ala Leu Gly Leu Arg Cys Gly Glu Ala Val Thr
                      20 25 30
          Leu Leu Asp Arg Gln Asp Trp Arg Asp Gly Ala Pro Ala His Thr Ala
                  35 40 45
          Cys Ala Leu Glu Gly Leu Asn Leu Val Pro Glu Val Lys Ile Leu Ala
              50 55 60
          Glu Pro Gly Phe Ala Lys Arg His Leu Pro Arg Gln Asn Thr Ala Lys
          65 70 75 80
          Ala Leu Met Ile Lys Tyr Glu Gln Arg Gln Gly Leu Met Ala Arg Asp
                          85 90 95
          Trp His Asp Trp Glu Arg Arg Cys Ala Pro Val Leu Asn Leu Leu Gly
                      100 105 110
          Leu His Phe Ala Thr Asp Gly Tyr Thr Pro Val Arg Arg Gly Pro Ala
                  115 120 125
          Arg Asp Phe Leu Ala Trp Gly Tyr Phe Gln Ser Glu Ala Tyr Phe Ala
              130 135 140
          Asp Phe Ala Pro Thr Ile Arg Ala Glu Leu Arg Ala Lys Gln Ala Pro
          145 150 155 160
          Ala Gly Val Trp Ala Glu Lys Ile Arg Ala Ala Ala Cys Pro Val Ala
                          165 170 175
          Leu His Leu Arg Arg Gly Asp Tyr Cys Arg Pro Glu Asn Glu Ile Leu
                      180 185 190
          Gln Val Cys Ser Pro Ala Tyr Tyr Ala Arg Ala Ala Ala Ala Ala Ala
                  195 200 205
          Ala Ala Tyr Pro Glu Ala Thr Leu Phe Val Phe Ser Asp Asp Ile Asp
              210 215 220
          Trp Ala Lys Glu His Leu Asp Thr Ala Gly Leu Pro Ala Val Trp Met
          225 230 235 240
          Pro Arg Gly Asp Ala Val Gly Asp Leu Asn Leu Met Ala Leu Cys Arg
                          245 250 255
          Gly Phe Ile Leu Ser Asn Ser Thr Tyr Ser Trp Trp Ala Gln Tyr Leu
                      260 265 270
          Ala Gly Glu Gly Arg Thr Val Trp Ala Pro Asp Arg Trp Phe Ala His
                  275 280 285
          Thr Lys Gln Thr Ala Leu Tyr Gln Pro Gly Trp His Leu Ile Glu Thr
              290 295 300
          Arg
          305
           <![CDATA[ <210> 82]]>
           <![CDATA[ <211> 288]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Pacific Oyster]]>
           <![CDATA[ <400> 82]]>
          Met Tyr Glu Ser Lys Lys Lys Leu Asp Gly Ile Ala Cys Val Lys Phe Gln
          1 5 10 15
          Gly Arg Leu Gly Asn Leu Met Met Glu Tyr Val Phe Leu Tyr Val Ile
                      20 25 30
          Ala Lys Met Lys His Leu Tyr Pro Ile Val Pro Glu Asn Ser Glu Leu
                  35 40 45
          Phe Gln Ile Phe Asp Leu Glu Lys Thr Thr Leu Ser Ala Ile Asn Lys
              50 55 60
          Ser Thr Asp Ser Cys Ser Lys Leu Pro Glu Tyr Lys Glu Ser Trp Gly
          65 70 75 80
          Leu Ser Tyr Asp Glu Lys Leu Leu Ala Val Pro Pro Asn Lys Ser Val
                          85 90 95
          Arg Phe Asn Gly Tyr Phe Gln Ser Trp Lys Tyr Trp Ile Lys Tyr Glu
                      100 105 110
          Asp Lys Ile Arg Lys Leu Leu Arg Phe Lys Asn Pro Ile Arg Gln Lys
                  115 120 125
          Ala His Asp Gln Met Arg Thr Ile Met Asn Lys Met Lys Phe Glu Val
              130 135 140
          Asn Lys Asp Ser Ala Ile Val Ser Ile His Ile Arg Arg Gly Asp Tyr
          145 150 155 160
          Ala Thr Glu Gly His Tyr Lys Tyr Gly Lys Leu Thr Pro Asn Glu Thr
                          165 170 175
          Tyr Tyr Ala Asn Ala Leu Gln Tyr Phe Lys Thr Arg His Lys Asn Ile
                      180 185 190
          Leu Phe Val Val Gly Ser Asn Asp Ile Asp Trp Ser Lys Lys Ala Leu
                  195 200 205
          Ala Lys Glu Lys Asn Val Tyr Tyr Ser Thr Gly Asn Ser Pro Ala Glu
              210 215 220
          Asp Ile Ala Leu Leu Ser Leu Ala Asn His Thr Ile Met Ser Val Gly
          225 230 235 240
          Thr Phe Gly Trp Trp Ile Gly Trp Met Ala Gln Gly Thr Ser Ile Phe
                          245 250 255
          Tyr Lys Asn Ile Phe Lys Ser Gln Ser Asp Phe Ala Lys Glu Phe Arg
                      260 265 270
          Asn Asn Ser Thr Asp Asp Phe Ile Tyr Pro Gly Trp Ile Pro Met Glu
                  275 280 285
           <![CDATA[ <210> 83]]>
           <![CDATA[ <211> 265]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Pacific Oyster]]>
           <![CDATA[ <400> 83]]>
          Met Glu Tyr Val Phe Leu Tyr Val Ile Ala Lys Met Lys His Leu Tyr
          1 5 10 15
          Pro Val Val Pro Glu Asn Ser Glu Leu Phe Gln Ile Phe Asn Ile Glu
                      20 25 30
          Lys Thr Thr Leu Thr Ala Ile Gly Lys Ser Ala Tyr Ser Cys Ala Lys
                  35 40 45
          Leu Pro Val Tyr Lys Glu Arg Trp Gly Leu Ser Tyr Asp Glu Lys Leu
              50 55 60
          His Ala Ile Pro Pro Tyr Lys Gly Val Gln Phe Asp Gly Tyr Phe Gln
          65 70 75 80
          Ser Trp Lys Tyr Trp Ile Lys Tyr Glu Asn Glu Ile Arg Lys Leu Leu
                          85 90 95
          Arg Phe Lys Asn Pro Ile Thr Gln Lys Ala Phe Thr Gln Met Arg Asp
                      100 105 110
          Ile Ile Asp Lys Met Lys Phe Glu Val Asn Lys Glu Ser Val Ile Val
                  115 120 125
          Ser Ile His Ile Arg Arg Gly Asp Tyr Ala Thr Glu Gly His Tyr Lys
              130 135 140
          Tyr Gly Lys Leu Thr Pro Asn Glu Thr Tyr Tyr Ala Asn Ala Met Gln
          145 150 155 160
          Tyr Phe Lys Thr Arg His Lys Asn Ile Leu Phe Val Val Gly Ser Asn
                          165 170 175
          Asp Ile Asp Trp Ser Lys Lys Lys Ala Leu Ala Arg Glu Lys Asn Val Tyr
                      180 185 190
          Tyr Ser Thr Gly Asn Ser Pro Ala Glu Asp Ile Ala Leu Leu Ser Leu
                  195 200 205
          Ala Asn His Thr Ile Met Ser Val Gly Thr Phe Gly Trp Trp Ile Gly
              210 215 220
          Trp Met Ala Gln Gly Thr Ser Val Phe Tyr Lys Asn Ile Phe Arg Pro
          225 230 235 240
          Gln Ser Asp Phe Ala Lys Glu Phe Arg Asn Asn Ser Ile Asp Asp Phe
                          245 250 255
          Ile Tyr Pro Gly Trp Ile Pro Met Glu
                      260 265
           <![CDATA[ <210> 84]]>
           <![CDATA[ <211> 277]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Uncultivated bacteria]]>
           <![CDATA[ <400> 84]]>
          Met Leu Thr Leu Lys Leu Lys Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Ala Ser His Asn Leu Ala Lys Asn Lys Lys Thr Lys Ile Asn
                      20 25 30
          Phe Asp Leu Ser Phe Phe Ser Asp Ile Glu Val Arg Asp Ile Lys Arg
                  35 40 45
          Asp Tyr Leu Leu Asp Lys Phe Asn Ile Ser Ala Asp Ile Ser Phe Asp
              50 55 60
          Gln Lys Asn Ser Ile Ser Gly Phe Arg Lys Phe Leu Val Lys Val Ile
          65 70 75 80
          Ser Lys Phe Phe Gly Glu Val Phe Tyr Tyr Arg Leu Lys Phe Leu Ser
                          85 90 95
          Ser Lys Tyr Leu Asp Gly Tyr Phe Gln Ser Glu Lys Tyr Phe Lys Asn
                      100 105 110
          Val Glu Glu Asp Ile Arg Lys Asp Phe Thr Leu Lys Asp Glu Met Gly
                  115 120 125
          Val Glu Ala Lys Lys Ile Glu Gln Gln Ile Val Asn Ser Lys Asn Ser
              130 135 140
          Val Ser Leu His Ile Arg Arg Gly Asp Tyr Val Asp Asp Leu Lys Thr
          145 150 155 160
          Asn Ile Tyr His Gly Val Cys Asn Leu Asp Tyr Tyr Lys Arg Ser Ile
                          165 170 175
          Lys Tyr Leu Lys Glu Asn Phe Gly Glu Ile Asn Ile Phe Val Phe Ser
                      180 185 190
          Asp Asp Ile Ala Trp Val Lys Glu Asn Leu Ala Phe Glu Asn Leu Gln
                  195 200 205
          Phe Val Ser Arg Pro Asp Ile Lys Asp Tyr Glu Glu Leu Met Leu Met
              210 215 220
          Ser Lys Cys Glu His Asn Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp
          225 230 235 240
          Gly Ala Trp Leu Asn Glu Asn Lys Asn Lys Ile Ile Ile Ala Pro Lys
                          245 250 255
          Glu Trp Phe Gln Lys Phe Asn Ile Asn Glu Lys His Ile Val Pro Lys
                      260 265 270
          Ser Trp Ile Arg Leu
                  275
           <![CDATA[ <210> 85]]>
           <![CDATA[ <211> 293]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Clostridium sp. CAG:510]]>
           <![CDATA[ <400> 85]]>
          Met Ile Met Leu Gln Met Thr Gly Gly Met Gly Asn Gln Met Phe Thr
          1 5 10 15
          Tyr Ala Leu Tyr Arg Ser Leu Arg Gln Lys Gly Lys Glu Val Cys Ile
                      20 25 30
          Glu Asp Phe Thr His Tyr Asp Thr Pro Glu Lys Asn Cys Leu Gln Thr
                  35 40 45
          Val Phe His Leu Asp Tyr Arg Lys Ala Asp Arg Glu Val Tyr Gln Arg
              50 55 60
          Leu Thr Asp Ser Glu Pro Asp Phe Leu His Lys Val Lys Arg Lys Leu
          65 70 75 80
          Thr Gly Arg Lys Glu Lys Ile Tyr Gln Glu Lys Asp Ala Ile Ile Phe
                          85 90 95
          Glu Pro Glu Val Phe Gln Thr Asp Asp Val Tyr Met Ile Gly Tyr Phe
                      100 105 110
          Gln Ser Gly Arg Tyr Phe Glu Lys Ala Val Phe Asp Leu Arg Lys Asp
                  115 120 125
          Phe Thr Phe Ala Trp Asn Thr Phe Pro Glu Lys Ala Lys Lys Leu Arg
              130 135 140
          Glu Gln Met Gln Ala Glu Ser Ser Val Ser Leu His Ile Arg Arg Gly
          145 150 155 160
          Asp Tyr Met Asn Gly Lys Phe Ala Ser Ile Tyr Gly Asn Ile Cys Thr
                          165 170 175
          Asp Ala Tyr Tyr Glu Ala Ala Arg Arg Tyr Met Lys Glu His Phe Gly
                      180 185 190
          Asp Cys Arg Phe Tyr Leu Phe Thr Asp Asp Ala Glu Trp Gly Arg Gln
                  195 200 205
          Gln Glu Ser Glu Asp Thr Val Tyr Val Asp Ala Ser Glu Gly Ala Gly
              210 215 220
          Ala Tyr Val Asp Met Ala Leu Met Ser Cys Cys Arg His His Ile Ile
          225 230 235 240
          Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala Trp Leu Asp Glu Asn Pro
                          245 250 255
          Asp Lys Thr Val Ile Ala Pro Ala Lys Trp Leu Asn Ile Ser Glu Gly
                      260 265 270
          Lys Asp Ile Tyr Ala Gly Leu Cys Asn Cys Leu Ile Asp Ala Asn Gly
                  275 280 285
          Ser Val Gln Gly Glu
              290
           <![CDATA[ <210> 86]]>
           <![CDATA[ <211> 351]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Robustazer Leech]]>
           <![CDATA[ <400> 86]]>
          Met Ser Phe Ile Ile Lys Tyr Val Tyr Val Val Ala Leu Ala Thr Ile
          1 5 10 15
          Leu Ile Phe Leu Ile Thr Val Tyr Phe Ala Glu Asn Phe Lys Val Gly
                      20 25 30
          Arg Asn Phe Leu Asp Ser Ser Pro Val Thr Pro Pro Gly Ser Lys Tyr
                  35 40 45
          Ala Ser Leu Asp Ala Cys Thr Phe Asn Ser Arg Arg Thr Gly Asn Leu
              50 55 60
          Met Phe Ala Leu Ala Gly Leu Leu Phe Val Ala Gln Asn Thr Arg Arg
          65 70 75 80
          Asn Pro Ile Leu Pro Thr Asn Ile Pro Tyr Gly Trp Met Asp Asp Tyr
                          85 90 95
          Phe Asn Thr Ser Ser Ile Gln Arg Leu Pro Asn Glu Phe Val Tyr Asp
                      100 105 110
          Ala Asn Lys Thr Val Val Leu Lys Glu Arg Phe Gly Pro Phe Val Tyr
                  115 120 125
          Asp Pro Ile Phe Glu Asn Pro His Lys Asn Ser Thr Val Glu Ser Lys
              130 135 140
          Glu Ile Ile Leu Leu Cys Gly Tyr Phe Gln Asn Tyr Lys Tyr Val Glu
          145 150 155 160
          Lys Ile Asp Lys Lys Leu Lys Met Ile Phe Thr Phe Tyr Asn Glu Thr
                          165 170 175
          Gln Ser Lys Val Gln Asn Phe Ile Asp His His Lys Lys Thr Ser Ala
                      180 185 190
          Lys Asn Arg Ser Asn Val Ala Thr Val Gly Ile His Ile Arg Arg Gly
                  195 200 205
          Asp Phe Leu Ile Lys Phe His Arg Asn Arg Gly Phe Ala Val Val Asp
              210 215 220
          Glu Asn Phe Ile Asn Ser Thr Val Asn Tyr Phe Tyr Lys Leu Trp Thr
          225 230 235 240
          Ala Lys Asn Val Glu Phe Ile Leu Tyr Phe Ile Ala Ser Glu Asp Glu
                          245 250 255
          Ala Trp Val Asn Ser Val Val Lys Lys Leu Asn Phe Ala Arg Ala Leu
                      260 265 270
          Asn Lys Ala Ala Phe Val Phe Ser Thr Ser Asn Gly Gly Pro Phe Asp
                  275 280 285
          Met Cys Leu Ile Ser Met Cys Asp Gly Val Ile Thr Ser Ser Gly Ser
              290 295 300
          Phe Ser Trp Trp Ala Gly Trp Leu Ala Asn Thr Thr Thr Ile Tyr Phe
          305 310 315 320
          Thr Gly Tyr Pro Arg Lys Gly Ser Ala Leu Gly Glu Gly Phe Asp Arg
                          325 330 335
          Lys Thr Tyr Gln Lys Pro Asp Trp Ile Gly Phe Pro Pro Glu Ile
                      340 345 350
           <![CDATA[ <210> 87]]>
           <![CDATA[ <211> 297]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Escherichia coli]]>
           <![CDATA[ <400> 87]]>
          Met Ser Ile Ile Arg Leu Gln Gly Gly Leu Gly Asn Gln Leu Phe Gln
          1 5 10 15
          Phe Ser Phe Gly Tyr Ala Leu Ser Lys Ile Asn Gly Thr Pro Leu Tyr
                      20 25 30
          Phe Asp Ile Ser His Tyr Ala Glu Asn Asp Asp His Gly Gly Tyr Arg
                  35 40 45
          Leu Asn Asn Leu Gln Ile Pro Glu Glu Tyr Leu Gln Tyr Tyr Thr Pro
              50 55 60
          Lys Ile Asn Asn Ile Tyr Lys Phe Leu Val Arg Gly Ser Arg Leu Tyr
          65 70 75 80
          Pro Glu Ile Phe Leu Phe Leu Gly Phe Cys Asn Glu Phe His Ala Tyr
                          85 90 95
          Gly Tyr Asp Phe Glu Tyr Ile Ala Gln Lys Trp Lys Ser Lys Lys Tyr
                      100 105 110
          Ile Gly Tyr Trp Gln Ser Glu His Phe Phe His Lys His Ile Leu Asp
                  115 120 125
          Leu Lys Glu Phe Phe Ile Pro Lys Asn Val Ser Glu Gln Ala Asn Leu
              130 135 140
          Leu Ala Ala Lys Ile Leu Glu Ser Gln Ser Ser Leu Ser Ile His Ile
          145 150 155 160
          Arg Arg Gly Asp Tyr Ile Lys Asn Lys Thr Ala Thr Leu Thr His Gly
                          165 170 175
          Val Cys Ser Leu Glu Tyr Tyr Lys Lys Ala Leu Asn Lys Ile Arg Asp
                      180 185 190
          Leu Ala Met Ile Arg Asp Val Phe Ile Phe Ser Asp Asp Ile Phe Trp
                  195 200 205
          Cys Lys Glu Asn Ile Glu Thr Leu Leu Ser Lys Lys Lys Tyr Asn Ile Tyr
              210 215 220
          Tyr Ser Glu Asp Leu Ser Gln Glu Glu Asp Leu Trp Leu Met Ser Leu
          225 230 235 240
          Ala Asn His His Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala
                          245 250 255
          Tyr Leu Gly Thr Ser Ala Ser Gln Ile Val Ile Tyr Pro Thr Pro Trp
                      260 265 270
          Tyr Asp Ile Thr Pro Lys Asn Thr Tyr Ile Pro Ile Val Asn His Trp
                  275 280 285
          Ile Asn Val Asp Lys His Ser Ser Cys
              290 295
           <![CDATA[ <210> 88]]>
           <![CDATA[ <211> 280]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Francisella sp. FSC1006]]>
           <![CDATA[ <400> 88]]>
          Met Lys Val Val Lys Ile Gln Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Phe Tyr Met Ser Leu Lys Gln Lys Tyr Lys Asp Cys Cys Ile
                      20 25 30
          Asp Ile Arg Asp Phe Glu Thr Tyr Thr Gln His Asn Gly Phe Glu Leu
                  35 40 45
          Asp Arg Val Phe Glu Asn Ile Arg Gln Ser Ile Ser Leu Cys Arg Cys
              50 55 60
          Lys Arg Lys Phe Phe Arg Ser Leu Phe Ser Lys Phe Leu Asn Arg Phe
          65 70 75 80
          Ile Lys Tyr His Lys Asn Tyr Phe Ser Gln Asp Asp Phe Gly Phe Asn
                          85 90 95
          Lys Lys Tyr Tyr Asn Lys Asp Asn Cys Tyr Leu Asp Gly Tyr Trp Gln
                      100 105 110
          Ser Glu Lys Tyr Phe Lys Ser Val Glu Lys Gln Ile Arg Glu Ile Phe
                  115 120 125
          Lys Phe Gln Thr Leu Asp Asp Lys Asn Ala Lys Ile Leu Glu Glu Tyr
              130 135 140
          Lys Asn Arg Ser Leu Val Ser Ile His Val Arg Arg Gly Asp Tyr Ile
          145 150 155 160
          Asn His Pro Leu His Gly Asp Ile Cys Asn Leu Asp Tyr Tyr Asn Asn
                          165 170 175
          Ala Ile Asp Ile Ile Lys Ser Arg Val Glu Ser Pro His Phe Phe Val
                      180 185 190
          Phe Ser Asp Asp Ile Glu Trp Cys Lys Gln Cys Leu Asp Ile Glu Asp
                  195 200 205
          Val Thr Tyr Ile Cys Thr Asn Thr Gly Ser Asp Ser Tyr Arg Asp Met
              210 215 220
          Gln Ile Met Ser Ile Cys Lys His Asn Ile Ile Ala Asn Ser Ser Phe
          225 230 235 240
          Ser Trp Trp Gly Ala Trp Leu Asn Gln Asn Ser Glu Lys Ile Ile Ile
                          245 250 255
          Ala Pro Asn Arg Trp Phe Asn Asp Asp Ser Ile Asn Gln Ser Asp Ile
                      260 265 270
          Cys Pro Glu Ser Trp Ile Lys Ile
                  275 280
           <![CDATA[ <210> 89]]>
           <![CDATA[ <211> 290]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Radiation-resistant spirochetes]]>
           <![CDATA[ <400> 89]]>
          Met Ile Ile Ser Arg Ile Thr Ser Gly Leu Gly Asn Gln Leu Phe Gln
          1 5 10 15
          Tyr Ala Thr Gly Arg His Leu Ala Leu Lys Asn Lys Ala Ala Leu Tyr
                      20 25 30
          Leu Asp Leu Ser Tyr Tyr Leu Tyr Glu Tyr Ser Asp Asp Thr Val Arg
                  35 40 45
          Pro Phe Lys Leu Asp His Phe Thr Val Pro Tyr Arg Leu Leu Gln Lys
              50 55 60
          Ser Pro Met Glu Tyr Val Ser Lys Ala Thr Lys Leu Leu Pro Asn Arg
          65 70 75 80
          Ser Leu Pro Pro Val Val Arg Phe Leu Lys Glu Lys His Phe His Phe
                          85 90 95
          Asp Gln Ser Ile Leu Asp Ala Arg Ala Ala Cys Ile Ile Leu Glu Gly
                      100 105 110
          Phe Trp Gln Ser Glu Ala Tyr Phe Arg Asp Asp Ala Asp Thr Ile Arg
                  115 120 125
          Gln Glu Leu Thr Leu Ala Gly Val Pro Ser Pro Glu Phe Ser His Tyr
              130 135 140
          Gln Gln Gln Ile Arg Asn Thr Ala Thr Pro Val Ser Ile His Ile Arg
          145 150 155 160
          Arg Gly Asp Tyr Val Asn His Pro Glu Phe Ser Lys Thr Phe Gly Phe
                          165 170 175
          Val Gly Leu Asp Tyr Tyr Lys Gln Ala Leu Gln Gln Leu Asn Glu His
                      180 185 190
          Phe Asp Gln Thr Gln Leu Tyr Val Phe Ser Asp Asp His Ala Trp Val
                  195 200 205
          His Glu Asn Leu Thr Leu Pro Asp Asn Thr Val Phe Val Gln Asn Thr
              210 215 220
          Gly Pro Asn Gly Asp Val Ala Asp Leu Val Leu Met Ser Tyr Cys Arg
          225 230 235 240
          His His Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala Trp Leu
                          245 250 255
          Asn Pro His Ser Asp Lys Leu Val Ile Thr Pro Arg Lys Trp Tyr Lys
                      260 265 270
          Gln Gln Pro Thr Trp Asn Thr Lys Asp Leu Leu Pro Ser Ala Trp Leu
                  275 280 285
          Ala Leu
              290
           <![CDATA[ <210> 90]]>
           <![CDATA[ <211> 291]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Radiation-resistant spirochetes]]>
           <![CDATA[ <400> 90]]>
          Met Val Ile Ser Val Leu Ala Gly Gly Leu Gly Asn Gln Leu Phe Gln
          1 5 10 15
          Tyr Ala Phe Gly Phe Ser Leu Ala Gln Gln Arg Lys Thr Glu Leu Arg
                      20 25 30
          Leu Glu Arg His Leu Leu Glu Ser Arg Thr Leu Ala Arg Leu Arg Asn
                  35 40 45
          Tyr Thr Pro Arg Gln Tyr Glu Leu Ser Val Phe Gly Ile Asp Gln Leu
              50 55 60
          Asp Ser Ser Leu Tyr Asp Thr Leu Arg Cys Leu Ser Arg Thr Leu Leu
          65 70 75 80
          Pro Gly Gln Gln Ala Val Leu Leu Arg Glu Ser Ala Pro Ala Thr Leu
                          85 90 95
          Thr Ser Leu Thr Gln Ser Asp Thr Gln Leu Glu Asp Ala Phe Cys Val
                      100 105 110
          Gly Tyr Trp Gln Ser Glu Ser Tyr Phe Lys Pro Val Glu Val Ala Leu
                  115 120 125
          Arg Gln Gln Leu Thr Val Lys Lys Pro Leu Ser Asn Leu Thr Leu Arg
              130 135 140
          Met Ala Glu Thr Ile Phe Ser Ser Ala Asn Ala Thr Phe Val His Ile
          145 150 155 160
          Arg Arg Gly Asp Tyr Ile Thr Asn Thr Lys Ala Asn Arg His His Gly
                          165 170 175
          Val Cys Gly Glu Ala Tyr Tyr Arg Arg Ala Val Glu Tyr Leu His Gln
                      180 185 190
          Gln Ala Ser Asp Ile Gln Phe Phe Val Phe Ser Asp Asp Gln Ala Trp
                  195 200 205
          Val Lys Arg Glu Leu Gly Ala Leu Leu Ser Pro Ala Ile Phe Val Glu
              210 215 220
          His Asn Thr Gly Val Asp Ser Trp Gln Asp Met Tyr Leu Met Ser Leu
          225 230 235 240
          Cys Arg His Ala Ile Val Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala
                          245 250 255
          Trp Leu Asn Pro Asp Leu Asn Arg Ile Val Val Ala Pro Gln Gln Trp
                      260 265 270
          Phe Ala Val Asn Arg Leu Thr Gln Pro Ala Val Ala Pro Ser His Trp
                  275 280 285
          Ile Thr Val
              290
           <![CDATA[ <210> 91]]>
           <![CDATA[ <211> 292]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Flavobacterium johnsonii]]>
           <![CDATA[ <400> 91]]>
          Met Asp Val Ile Val Ile Phe Asn Gly Leu Gly Asn Gln Met Ser Gln
          1 5 10 15
          Tyr Ala Phe Tyr Leu Gln Lys Lys Lys Ile Asn Pro Ser Thr Tyr Leu
                      20 25 30
          Ile Ser Phe Cys Lys Asp His Asn Gly Leu Glu Ile Asn Ser Ile Phe
                  35 40 45
          Asn Ile Thr Trp Lys Asn Thr Leu Ile Asn Lys Ser Leu Tyr Phe Leu
              50 55 60
          Phe Arg Ile Leu Ile Ala Asp Arg Phe Lys Leu Ile Thr Val Pro Leu
          65 70 75 80
          Lys Lys Leu Leu Thr Gln Met Gly Cys Ser Val Ile Asn Glu Asp Tyr
                          85 90 95
          Asp Tyr Asn Phe Asn Ser Glu Tyr Leu Lys Pro Ser Lys Gly Ile Thr
                      100 105 110
          Phe Tyr Phe Gly Gly Trp His Asn Glu Lys Tyr Phe Ile Asp Ser Lys
                  115 120 125
          Asp Glu Ile Leu Asp Ser Phe Ser Phe Ser Lys Asp Val Asp Glu Ile
              130 135 140
          Thr Thr Ser Leu Val Gln Glu Met Ala Asn Tyr Glu Ser Val Ser Ile
          145 150 155 160
          His Val Arg Arg Gly Asp Tyr Leu Asn Ala Ala Asn Ile Asn Leu Phe
                          165 170 175
          Gly Lys Val Cys Thr Lys Glu Tyr Phe Glu Lys Ala Ile Asp Leu Met
                      180 185 190
          Asn Glu Lys Val Lys Thr Pro His Tyr Tyr Ile Phe Ser Asn Asp Ile
                  195 200 205
          Gly Trp Val Lys Glu Asn Phe Asn Leu Glu Arg Val Thr Tyr Val Thr
              210 215 220
          His Asn Ser Gly Ile Asn Ser Trp Arg Asp Met Phe Leu Met Ser Ser
          225 230 235 240
          Cys Lys His Asn Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala
                          245 250 255
          Trp Leu Asn Lys Asn Asn Asn Asn Thr Val Leu Ser Pro Thr Lys Phe
                      260 265 270
          Leu Asn Ser Asp Lys Val Ser Asp Ile Tyr Pro Ala Asn Trp Thr Lys
                  275 280 285
          Ile Ser Asp Asn
              290
           <![CDATA[ <210> 92]]>
           <![CDATA[ <211> 300]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Helicobacter pylori]]>
           <![CDATA[ <400> 92]]>
          Met Ala Phe Lys Val Val Gln Ile Cys Gly Gly Leu Gly Asn Gln Met
          1 5 10 15
          Phe Gln Tyr Ala Phe Ala Lys Ser Leu Gln Lys His Ser Asn Thr Pro
                      20 25 30
          Val Leu Leu Asp Ile Thr Ser Phe Asp Trp Ser Asp Arg Lys Met Gln
                  35 40 45
          Leu Glu Leu Phe Pro Ile Asp Leu Pro Tyr Ala Ser Ala Lys Glu Ile
              50 55 60
          Ala Ile Ala Lys Met Gln His Leu Pro Lys Leu Val Arg Asp Ala Leu
          65 70 75 80
          Lys Cys Met Gly Phe Asp Arg Val Ser Gln Glu Ile Val Phe Glu Tyr
                          85 90 95
          Glu Pro Lys Leu Leu Lys Pro Ser Arg Leu Thr Tyr Phe Phe Gly Tyr
                      100 105 110
          Phe Gln Asp Pro Arg Tyr Phe Asp Ala Ile Ser Pro Leu Ile Lys Gln
                  115 120 125
          Thr Phe Thr Leu Pro Pro Pro Pro Glu Asn Asn Lys Asn Asn Asn Lys
              130 135 140
          Lys Glu Glu Glu Tyr Gln Cys Lys Leu Ser Leu Ile Leu Ala Ala Lys
          145 150 155 160
          Asn Ser Val Phe Val His Ile Arg Arg Gly Asp Tyr Val Gly Ile Gly
                          165 170 175
          Cys Gln Leu Gly Ile Asp Tyr Gln Lys Lys Ala Leu Glu Tyr Met Ala
                      180 185 190
          Lys Arg Val Pro Asn Met Glu Leu Phe Val Phe Cys Glu Asp Leu Glu
                  195 200 205
          Phe Thr Gln Asn Leu Asp Leu Gly Tyr Pro Phe Met Asp Met Thr Thr
              210 215 220
          Arg Asp Lys Glu Glu Glu Ala Tyr Trp Asp Met Leu Leu Met Gln Ser
          225 230 235 240
          Cys Gln His Gly Ile Ile Ala Asn Ser Thr Tyr Ser Trp Trp Ala Ala
                          245 250 255
          Tyr Leu Ile Glu Asn Pro Glu Lys Ile Ile Ile Gly Pro Lys His Trp
                      260 265 270
          Leu Phe Gly His Glu Asn Ile Leu Cys Lys Glu Trp Val Lys Ile Glu
                  275 280 285
          Ser His Phe Glu Val Lys Ser Gln Lys Tyr Asn Ala
              290 295 300
           <![CDATA[ <210> 93]]>
           <![CDATA[ <211> 298]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Helicobacter pylori]]>
           <![CDATA[ <400> 93]]>
          Met Ala Phe Lys Val Val Gln Ile Cys Gly Gly Leu Gly Asn Gln Met
          1 5 10 15
          Phe Gln Tyr Ala Phe Ala Lys Ser Leu Gln Lys His Leu Asn Thr Pro
                      20 25 30
          Val Leu Leu Asp Ile Thr Ser Phe Asp Trp Ser Asn Arg Lys Met Gln
                  35 40 45
          Leu Glu Leu Phe Pro Ile Asp Leu Pro Tyr Ala Ser Ala Lys Glu Ile
              50 55 60
          Ala Ile Ala Lys Met Gln His Leu Pro Lys Leu Val Arg Asp Thr Leu
          65 70 75 80
          Lys Cys Met Gly Phe Asp Arg Val Ser Gln Glu Ile Val Phe Glu Tyr
                          85 90 95
          Glu Pro Gly Leu Leu Lys Pro Ser Arg Leu Thr Tyr Phe Tyr Gly Tyr
                      100 105 110
          Phe Gln Asp Pro Arg Tyr Phe Asp Ala Ile Ser Pro Leu Ile Lys Gln
                  115 120 125
          Thr Phe Thr Leu Pro Pro Pro Glu Asn Gly Asn Asn Lys Lys Lys Glu
              130 135 140
          Glu Glu Tyr His Arg Lys Leu Ala Leu Ile Leu Ala Ala Lys Asn Ser
          145 150 155 160
          Val Phe Val His Val Arg Arg Gly Asp Tyr Val Gly Ile Gly Cys Gln
                          165 170 175
          Leu Gly Ile Asp Tyr Gln Lys Lys Ala Leu Glu Tyr Ile Ala Lys Arg
                      180 185 190
          Val Pro Asn Met Glu Leu Phe Val Phe Cys Glu Asp Leu Lys Phe Thr
                  195 200 205
          Gln Asn Leu Asp Leu Gly Tyr Pro Phe Met Asp Met Thr Thr Arg Asp
              210 215 220
          Lys Glu Glu Glu Ala Tyr Trp Asp Met Leu Leu Met Gln Ser Cys Lys
          225 230 235 240
          His Gly Ile Ile Ala Asn Ser Thr Tyr Ser Trp Trp Ala Ala Tyr Leu
                          245 250 255
          Ile Asn Asn Pro Glu Lys Ile Ile Ile Gly Pro Lys His Trp Leu Phe
                      260 265 270
          Gly His Glu Asn Ile Leu Cys Lys Glu Trp Val Lys Ile Glu Ser His
                  275 280 285
          Phe Glu Val Lys Ser Lys Lys Tyr Asn Ala
              290 295
           <![CDATA[ <210> 94]]>
           <![CDATA[ <211> 286]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Helicobacter weasel]]>
           <![CDATA[ <400> 94]]>
          Met Asp Phe Lys Ile Val Gln Val His Gly Gly Leu Gly Asn Gln Met
          1 5 10 15
          Phe Gln Tyr Ala Phe Ala Lys Ser Leu Gln Thr His Leu Asn Ile Pro
                      20 25 30
          Val Leu Leu Asp Thr Thr Trp Phe Asp Tyr Gly Asn Arg Glu Leu Gly
                  35 40 45
          Leu His Leu Phe Pro Ile Asp Leu Gln Cys Ala Ser Ala Gln Gln Ile
              50 55 60
          Ala Ala Ala His Met Gln Asn Leu Pro Arg Leu Val Arg Gly Ala Leu
          65 70 75 80
          Arg Arg Met Gly Leu Gly Arg Val Ser Lys Glu Ile Val Phe Glu Tyr
                          85 90 95
          Met Pro Glu Leu Phe Glu Pro Ser Arg Ile Ala Tyr Phe His Gly Tyr
                      100 105 110
          Phe Gln Asp Pro Arg Tyr Phe Glu Asp Ile Ser Pro Leu Ile Lys Gln
                  115 120 125
          Thr Phe Thr Leu Pro His Pro Thr Glu His Ala Glu Gln Tyr Ser Arg
              130 135 140
          Lys Leu Ser Gln Ile Leu Ala Ala Lys Asn Ser Val Phe Val His Ile
          145 150 155 160
          Arg Arg Gly Asp Tyr Met Arg Leu Gly Trp Gln Leu Asp Ile Ser Tyr
                          165 170 175
          Gln Leu Arg Ala Ile Ala Tyr Met Ala Lys Arg Val Gln Asn Leu Glu
                      180 185 190
          Leu Phe Leu Phe Cys Glu Asp Leu Glu Phe Val Gln Asn Leu Asp Leu
                  195 200 205
          Gly Tyr Pro Phe Val Asp Met Thr Thr Arg Asp Gly Ala Ala His Trp
              210 215 220
          Asp Met Met Leu Met Gln Ser Cys Lys His Gly Ile Ile Thr Asn Ser
          225 230 235 240
          Thr Tyr Ser Trp Trp Ala Ala Tyr Leu Ile Lys Asn Pro Glu Lys Ile
                          245 250 255
          Ile Ile Gly Pro Ser His Trp Ile Tyr Gly Asn Glu Asn Ile Leu Cys
                      260 265 270
          Lys Asp Trp Val Lys Ile Glu Ser Gln Phe Glu Thr Lys Ser
                  275 280 285
           <![CDATA[ <210> 95]]>
           <![CDATA[ <211> 300]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Helicobacter pylori]]>
           <![CDATA[ <400> 95]]>
          Met Ala Phe Lys Val Val Gln Ile Cys Gly Gly Leu Gly Asn Gln Met
          1 5 10 15
          Phe Gln Tyr Ala Phe Ala Lys Ser Leu Gln Lys His Ser Asn Thr Pro
                      20 25 30
          Val Leu Leu Asp Ile Thr Ser Phe Asp Trp Ser Asp Arg Lys Met Gln
                  35 40 45
          Leu Glu Leu Phe Pro Ile Asn Leu Pro Tyr Ala Ser Ala Lys Glu Ile
              50 55 60
          Ala Ile Ala Lys Met Gln His Leu Pro Lys Leu Val Arg Asp Ala Leu
          65 70 75 80
          Lys Cys Met Gly Phe Asp Arg Val Ser Gln Glu Ile Val Phe Glu Tyr
                          85 90 95
          Glu Pro Glu Leu Leu Lys Pro Ser Arg Leu Thr Tyr Phe Tyr Gly Tyr
                      100 105 110
          Phe Gln Asp Pro Arg Tyr Phe Asp Ala Ile Ser Pro Leu Ile Lys Gln
                  115 120 125
          Thr Phe Thr Leu Pro Pro Pro Pro Glu Asn Asn Lys Asn Asn Asn Lys
              130 135 140
          Lys Glu Glu Glu Tyr His Arg Lys Leu Ser Leu Ile Leu Ala Ala Lys
          145 150 155 160
          Asn Ser Val Phe Val His Ile Arg Arg Gly Asp Tyr Val Gly Ile Gly
                          165 170 175
          Cys Gln Leu Gly Ile Asp Tyr Gln Lys Lys Ala Leu Glu Tyr Met Ala
                      180 185 190
          Lys Arg Val Pro Asn Met Glu Leu Phe Val Phe Cys Glu Asp Leu Glu
                  195 200 205
          Phe Thr Gln Asn Leu Asp Leu Gly Tyr Pro Phe Met Asp Met Thr Thr
              210 215 220
          Arg Asn Lys Glu Glu Glu Ala Tyr Trp Asp Met Leu Leu Met Gln Ser
          225 230 235 240
          Cys Gln His Gly Ile Ile Ala Asn Ser Thr Tyr Ser Trp Trp Ala Ala
                          245 250 255
          Tyr Leu Ile Glu Asn Pro Glu Lys Ile Ile Ile Gly Pro Lys His Trp
                      260 265 270
          Leu Phe Gly His Glu Asn Ile Leu Cys Lys Glu Trp Val Lys Ile Glu
                  275 280 285
          Ser His Phe Glu Val Lys Ser Gln Lys Tyr Asn Ala
              290 295 300
           <![CDATA[ <210> 96]]>
           <![CDATA[ <211> 288]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Prevotella niger producing]]>
           <![CDATA[ <400> 96]]>
          Met Lys Ile Val Lys Ile Leu Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Leu Tyr Leu Ser Leu Gln Glu Ser Phe Pro Lys Glu Arg Val
                      20 25 30
          Ala Leu Asp Leu Ser Ser Phe His Gly Tyr His Leu His Asn Gly Phe
                  35 40 45
          Glu Leu Glu Asn Ile Phe Ser Val Thr Ala Gln Lys Ala Ser Ala Ala
              50 55 60
          Asp Ile Met Arg Ile Ala Tyr Tyr Tyr Pro Asn Tyr Leu Leu Trp Arg
          65 70 75 80
          Ile Gly Lys Arg Phe Leu Pro Arg Arg Lys Gly Met Cys Leu Glu Ser
                          85 90 95
          Ser Ser Leu Arg Phe Asp Glu Ser Val Leu Arg Gln Glu Gly Asn Arg
                      100 105 110
          Tyr Phe Asp Gly Tyr Trp Gln Asp Glu Arg Tyr Phe Ala Ala Tyr Arg
                  115 120 125
          Glu Lys Val Leu Lys Ala Phe Thr Phe Pro Ala Phe Lys Arg Ala Glu
              130 135 140
          Asn Leu Ser Leu Leu Glu Lys Leu Asp Glu Asn Ser Ile Ala Leu His
          145 150 155 160
          Val Arg Arg Gly Asp Tyr Val Gly Asn Asn Leu Tyr Gln Gly Ile Cys
                          165 170 175
          Asp Leu Asp Tyr Tyr Arg Thr Ala Ile Glu Lys Met Cys Ala His Val
                      180 185 190
          Thr Pro Ser Leu Phe Cys Ile Phe Ser Asn Asp Ile Thr Trp Cys Gln
                  195 200 205
          Gln His Leu Gln Pro Tyr Leu Lys Ala Pro Val Val Tyr Val Thr Trp
              210 215 220
          Asn Thr Gly Val Glu Ser Tyr Arg Asp Met Gln Leu Met Ser Cys Cys
          225 230 235 240
          Ala His Asn Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala Trp
                          245 250 255
          Leu Asn Gln Asn Arg Glu Lys Val Val Ile Ala Pro Lys Lys Trp Leu
                      260 265 270
          Asn Met Glu Glu Cys His Phe Thr Leu Pro Ala Ser Trp Ile Lys Ile
                  275 280 285
           <![CDATA[ <210> 97]]>
           <![CDATA[ <211> 278]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Clostridium baumannii]]>
           <![CDATA[ <400> 97]]>
          Met Val Ile Ile Lys Met Met Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Leu Tyr Lys Ala Phe Glu Gln Lys His Ile Asp Val Tyr Ala
                      20 25 30
          Asp Leu Ala Trp Tyr Lys Asn Lys Ser Val Lys Phe Glu Leu Tyr Asn
                  35 40 45
          Phe Gly Ile Lys Ile Asn Val Ala Ser Glu Lys Asp Ile Asn Arg Leu
              50 55 60
          Ser Asp Cys Gln Ala Asp Phe Val Ser Arg Ile Arg Arg Lys Ile Phe
          65 70 75 80
          Gly Lys Lys Lys Ser Phe Val Ser Glu Lys Asn Asp Ser Cys Tyr Glu
                          85 90 95
          Asn Asp Ile Leu Arg Met Asp Asn Val Tyr Leu Ser Gly Tyr Trp Gln
                      100 105 110
          Thr Glu Lys Tyr Phe Ser Asn Thr Arg Glu Lys Leu Leu Glu Asp Tyr
                  115 120 125
          Ser Phe Ala Leu Val Asn Ser Gln Val Ser Glu Trp Glu Asp Ser Ile
              130 135 140
          Arg Asn Lys Asn Ser Val Ser Ile His Ile Arg Arg Gly Asp Tyr Leu
          145 150 155 160
          Gln Gly Glu Leu Tyr Gly Gly Ile Cys Thr Ser Leu Tyr Tyr Ala Glu
                          165 170 175
          Ala Ile Glu Tyr Ile Lys Met Arg Val Pro Asn Ala Lys Phe Phe Val
                      180 185 190
          Phe Ser Asp Asp Val Glu Trp Val Lys Gln Gln Glu Asp Phe Lys Gly
                  195 200 205
          Phe Val Ile Val Asp Arg Asn Glu Tyr Ser Ser Ala Leu Ser Asp Met
              210 215 220
          Tyr Leu Met Ser Leu Cys Lys His Asn Ile Ile Ala Asn Ser Ser Phe
          225 230 235 240
          Ser Trp Trp Ala Ala Trp Leu Asn Arg Asn Glu Glu Lys Ile Val Ile
                          245 250 255
          Ala Pro Arg Arg Trp Leu Asn Gly Lys Cys Thr Pro Asp Ile Trp Cys
                      260 265 270
          Lys Lys Trp Ile Arg Ile
                  275
           <![CDATA[ <210> 98]]>
           <![CDATA[ <211> 289]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Clostridium bacteria VE202-29]]>
           <![CDATA[ <400> 98]]>
          Met Val Ile Val Gln Leu Ser Gly Gly Leu Gly Asn Gln Met Phe Glu
          1 5 10 15
          Tyr Ala Leu Tyr Leu Ser Leu Lys Ala Lys Gly Lys Glu Val Lys Ile
                      20 25 30
          Asp Asp Val Thr Cys Tyr Glu Gly Pro Gly Thr Arg Pro Arg Gln Leu
                  35 40 45
          Asp Val Phe Gly Ile Thr Tyr Asp Arg Ala Ser Arg Glu Glu Leu Thr
              50 55 60
          Glu Met Thr Asp Ala Ser Met Asp Ala Leu Ser Arg Val Arg Arg Lys
          65 70 75 80
          Leu Thr Gly Arg Arg Thr Lys Ala Tyr Arg Glu Arg Asp Ile Asn Phe
                          85 90 95
          Asp Pro Leu Val Met Glu Lys Asp Pro Ala Leu Leu Glu Gly Cys Phe
                      100 105 110
          Gln Ser Asp Lys Tyr Phe Arg Asp Cys Glu Gly Arg Val Arg Glu Ala
                  115 120 125
          Tyr Arg Phe Arg Gly Ile Glu Ser Gly Ala Phe Pro Leu Pro Glu Asp
              130 135 140
          Tyr Leu Arg Leu Glu Lys Gln Ile Glu Asp Cys Gln Ser Val Ser Val
          145 150 155 160
          His Ile Arg Arg Gly Asp Tyr Leu Asp Glu Ser His Gly Gly Leu Tyr
                          165 170 175
          Thr Gly Ile Cys Thr Glu Ala Tyr Tyr Lys Glu Ala Phe Ala Arg Met
                      180 185 190
          Glu Arg Leu Val Pro Gly Ala Arg Phe Phe Leu Phe Ser Asn Asp Pro
                  195 200 205
          Glu Trp Thr Arg Glu His Phe Glu Ser Lys Asn Cys Val Leu Val Glu
              210 215 220
          Gly Ser Thr Glu Asp Thr Gly Tyr Met Asp Leu Tyr Leu Met Ser Arg
          225 230 235 240
          Cys Arg His Asn Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Gly Ala
                          245 250 255
          Trp Leu Asn Glu Asn Pro Glu Lys Lys Val Ile Ala Pro Ala Lys Trp
                      260 265 270
          Leu Asn Gly Arg Glu Cys Arg Asp Ile Tyr Thr Glu Arg Met Ile Arg
                  275 280 285
          Leu
           <![CDATA[ <210> 99]]>
           <![CDATA[ <211> 295]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Candidate swamp methanogens E1-9c]]>
           <![CDATA[ <400> 99]]>
          Met Ile Ile Val Arg Leu Lys Gly Gly Leu Gly Asn Gln Leu Ser Gln
          1 5 10 15
          Tyr Ala Leu Gly Arg Lys Ile Ala His Leu His Asn Thr Glu Leu Lys
                      20 25 30
          Leu Asp Thr Thr Trp Phe Thr Thr Ile Ser Ser Asp Thr Pro Arg Thr
                  35 40 45
          Tyr Arg Leu Asn Asn Tyr Asn Ile Ile Gly Thr Ile Ala Ser Ala Lys
              50 55 60
          Glu Ile Gln Leu Ile Glu Arg Gly Arg Ala Gln Gly Arg Gly Tyr Leu
          65 70 75 80
          Leu Ser Lys Ile Ser Asp Leu Leu Thr Pro Met Tyr Arg Arg Thr Tyr
                          85 90 95
          Val Arg Glu Arg Met His Thr Phe Asp Lys Ala Ile Leu Thr Val Pro
                      100 105 110
          Asp Asn Val Tyr Leu Asp Gly Tyr Trp Gln Thr Glu Lys Tyr Phe Lys
                  115 120 125
          Asp Ile Glu Glu Ile Leu Arg Arg Glu Val Thr Leu Lys Asp Glu Pro
              130 135 140
          Asp Ser Ile Asn Leu Glu Met Ala Glu Arg Ile Gln Ala Cys His Ser
          145 150 155 160
          Val Ser Leu His Val Arg Arg Gly Asp Tyr Val Ser Asn Pro Thr Thr
                          165 170 175
          Gln Gln Phe His Gly Cys Cys Ser Ile Asp Tyr Tyr Asn Arg Ala Ile
                      180 185 190
          Ser Leu Ile Glu Glu Lys Val Asp Asp Pro Ser Phe Phe Ile Phe Ser
                  195 200 205
          Asp Asp Leu Pro Trp Ala Lys Glu Asn Leu Asp Ile Pro Gly Glu Lys
              210 215 220
          Thr Phe Val Ala His Asn Gly Pro Glu Lys Glu Tyr Cys Asp Leu Trp
          225 230 235 240
          Leu Met Ser Leu Cys Gln His His Ile Ile Ala Asn Ser Ser Phe Ser
                          245 250 255
          Trp Trp Gly Ala Trp Leu Gly Gln Asp Ala Glu Lys Met Val Ile Ala
                      260 265 270
          Pro Arg Arg Trp Ala Leu Ser Glu Ser Phe Asp Thr Ser Asp Ile Ile
                  275 280 285
          Pro Asp Ser Trp Ile Thr Ile
              290 295
           <![CDATA[ <210> 100]]>
           <![CDATA[ <211> 287]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Tannerella sp. CAG:118]]>
           <![CDATA[ <400> 100]]>
          Met Val Arg Ile Val Glu Ile Ile Gly Gly Leu Gly Asn Gln Met Phe
          1 5 10 15
          Gln Tyr Ala Phe Ser Leu Tyr Leu Lys Asn Lys Ser His Ile Trp Asp
                      20 25 30
          Arg Leu Tyr Val Asp Ile Glu Ala Met Lys Thr Tyr Asp Arg His Tyr
                  35 40 45
          Gly Leu Glu Leu Glu Lys Val Phe Asn Leu Ser Leu Cys Pro Ile Ser
              50 55 60
          Asn Arg Leu His Arg Asn Leu Gln Lys Arg Ser Phe Ala Lys His Phe
          65 70 75 80
          Val Lys Ser Leu Tyr Glu His Ser Glu Cys Glu Phe Asp Glu Pro Val
                          85 90 95
          Tyr Arg Gly Leu Arg Pro Tyr Arg Tyr Tyr Arg Gly Tyr Trp Gln Asn
                      100 105 110
          Glu Gly Tyr Phe Val Asp Ile Glu Pro Met Ile Arg Glu Ala Phe Gln
                  115 120 125
          Phe Asn Val Asn Ile Leu Ser Lys Lys Thr Lys Ala Ile Ala Ser Lys
              130 135 140
          Met Arg Arg Glu Leu Ser Val Ser Ile His Val Arg Arg Gly Asp Tyr
          145 150 155 160
          Glu Asn Leu Pro Glu Ala Lys Ala Met His Gly Gly Ile Cys Ser Leu
                          165 170 175
          Asp Tyr Tyr His Lys Ala Ile Asp Phe Ile Arg Gln Arg Leu Asp Asn
                      180 185 190
          Asn Ile Cys Phe Tyr Leu Phe Ser Asp Asp Ile Asn Trp Val Glu Glu
                  195 200 205
          Asn Leu Gln Leu Glu Asn Arg Cys Ile Ile Asp Trp Asn Gln Gly Glu
              210 215 220
          Asp Ser Trp Gln Asp Met Tyr Leu Met Ser Cys Cys Arg His His Ile
          225 230 235 240
          Ile Ala Asn Ser Ser Phe Ser Trp Trp Ala Ala Trp Leu Asn Pro Asn
                          245 250 255
          Lys Asn Lys Ile Val Leu Thr Pro Asn Lys Trp Phe Asn His Thr Asp
                      260 265 270
          Ala Val Gly Ile Val Pro Lys Ser Trp Ile Lys Ile Pro Val Phe
                  275 280 285
           <![CDATA[ <210> 101]]>
           <![CDATA[ <211> 289]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Bacteroides faecalis]]>
           <![CDATA[ <400> 101]]>
          Met Lys Ile Val Lys Ile Leu Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Leu Phe Leu Ser Leu Lys Glu Arg Phe Pro His Glu Gln Val
                      20 25 30
          Met Ile Asp Thr Ser Cys Phe Arg Asn Tyr Pro Leu His Asn Gly Phe
                  35 40 45
          Glu Val Asp Arg Ile Phe Ala Gln Lys Ala Pro Val Ala Ser Trp Arg
              50 55 60
          Asn Ile Leu Lys Val Ala Tyr Pro Tyr Pro Asn Tyr Arg Phe Trp Lys
          65 70 75 80
          Ile Gly Lys Tyr Ile Leu Pro Lys Arg Lys Thr Met Cys Val Glu Arg
                          85 90 95
          Lys Asn Phe Ser Phe Asp Ala Ala Val Leu Thr Arg Lys Gly Asp Cys
                      100 105 110
          Tyr Tyr Asp Gly Tyr Trp Gln His Glu Glu Tyr Phe Cys Asp Met Lys
                  115 120 125
          Glu Thr Ile Trp Glu Ala Phe Ser Phe Pro Glu Pro Val Asp Gly Arg
              130 135 140
          Asn Lys Glu Ile Gly Ala Leu Leu Gln Ala Ser Asp Ser Ala Ser Leu
          145 150 155 160
          His Val Arg Arg Gly Asp Tyr Val Asn His Pro Leu Phe Arg Gly Ile
                          165 170 175
          Cys Asp Leu Asp Tyr Tyr Lys Arg Ala Ile His Tyr Met Glu Glu Arg
                      180 185 190
          Val Asn Pro Gln Leu Tyr Cys Val Phe Ser Asn Asp Met Ala Trp Cys
                  195 200 205
          Glu Ser His Leu Arg Ala Leu Leu Pro Gly Lys Glu Val Val Tyr Val
              210 215 220
          Asp Trp Asn Lys Gly Ala Glu Ser Tyr Val Asp Met Arg Leu Met Ser
          225 230 235 240
          Leu Cys Arg His Asn Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Gly
                          245 250 255
          Ala Trp Leu Asn Arg Asn Pro Gln Lys Val Val Val Ala Pro Glu Arg
                      260 265 270
          Trp Met Asn Ser Pro Ile Glu Asp Pro Val Ser Asp Lys Trp Ile Lys
                  275 280 285
          Leu
           <![CDATA[ <210> 102]]>
           <![CDATA[ <211> 290]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Vibrio butyricum species]]>
           <![CDATA[ <400> 102]]>
          Met Ile Ile Ile Gln Leu Lys Gly Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Leu Tyr Lys Ser Leu Lys Lys Arg Gly Lys Glu Val Lys Ile
                      20 25 30
          Asp Asp Lys Thr Gly Phe Val Asn Asp Lys Leu Arg Ile Pro Val Leu
                  35 40 45
          Ser Arg Trp Gly Val Glu Tyr Asp Arg Ala Thr Asp Glu Glu Ile Ile
              50 55 60
          Asn Leu Thr Asp Ser Lys Met Asp Leu Phe Ser Arg Ile Arg Arg Lys
          65 70 75 80
          Leu Thr Gly Arg Lys Thr Phe Arg Ile Asp Glu Glu Ser Gly Lys Phe
                          85 90 95
          Asn Pro Glu Ile Leu Glu Lys Glu Asn Ala Tyr Leu Val Gly Tyr Trp
                      100 105 110
          Gln Cys Asp Lys Tyr Phe Asp Asp Lys Asp Val Val Arg Glu Ile Arg
                  115 120 125
          Glu Ala Phe Glu Lys Lys Pro Gln Glu Leu Met Thr Asp Ala Ser Ser
              130 135 140
          Trp Ser Thr Leu Gln Gln Ile Glu Cys Cys Glu Ser Val Ser Leu His
          145 150 155 160
          Val Arg Arg Thr Asp Tyr Val Asp Glu Glu His Ile His Ile His Asn
                          165 170 175
          Ile Cys Thr Glu Lys Tyr Tyr Lys Asn Ala Ile Asp Arg Val Arg Lys
                      180 185 190
          Gln Tyr Pro Ser Ala Val Phe Phe Ile Phe Thr Asp Asp Lys Glu Trp
                  195 200 205
          Cys Arg Asp His Phe Lys Gly Pro Asn Phe Ile Val Val Glu Leu Glu
              210 215 220
          Glu Gly Asp Gly Thr Asp Ile Ala Glu Met Thr Leu Met Ser Arg Cys
          225 230 235 240
          Lys His His Ile Ile Ala Asn Ser Ser Phe Ser Trp Trp Ala Ala Trp
                          245 250 255
          Leu Asn Asp Ser Pro Glu Lys Ile Val Ile Ala Pro Gln Lys Trp Ile
                      260 265 270
          Asn Asn Arg Asp Met Asp Asp Ile Tyr Thr Glu Arg Met Thr Lys Ile
                  275 280 285
          Ala Leu
              290
           <![CDATA[ <210> 103]]>
           <![CDATA[ <211> 281]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Parabacter johnsonii]]>
           <![CDATA[ <400> 103]]>
          Met Arg Leu Ile Lys Met Ile Gly Gly Leu Gly Asn Gln Met Phe Ile
          1 5 10 15
          Tyr Ala Phe Tyr Leu Lys Met Lys His His Tyr Pro Asp Thr Asn Ile
                      20 25 30
          Asp Leu Ser Asp Met Val His Tyr Lys Val His Asn Gly Tyr Glu Met
                  35 40 45
          Asn Arg Ile Phe Asp Leu Ser Gln Thr Glu Phe Cys Ile Asn Arg Thr
              50 55 60
          Leu Lys Lys Ile Leu Glu Phe Leu Phe Phe Lys Lys Ile Tyr Glu Arg
          65 70 75 80
          Arg Gln Asp Pro Ser Thr Leu Tyr Pro Tyr Glu Lys Arg Tyr Phe Trp
                          85 90 95
          Pro Leu Leu Tyr Phe Lys Gly Phe Tyr Gln Ser Glu Arg Phe Phe Phe
                      100 105 110
          Asp Ile Lys Asp Asp Val Arg Lys Ala Phe Ser Phe Asn Leu Asn Ile
                  115 120 125
          Ala Asn Pro Glu Ser Leu Glu Leu Leu Lys Gln Ile Glu Val Asp Asp
              130 135 140
          Gln Ala Val Ser Ile His Ile Arg Arg Gly Asp Tyr Leu Leu Pro Arg
          145 150 155 160
          His Trp Ala Asn Thr Gly Ser Val Cys Gln Leu Pro Tyr Tyr Lys Asn
                          165 170 175
          Ala Ile Ala Glu Met Glu Asn Arg Ile Thr Gly Pro Ser Tyr Tyr Val
                      180 185 190
          Phe Ser Asp Asp Ile Ser Trp Val Lys Glu Asn Ile Pro Leu Lys Lys
                  195 200 205
          Ala Val Tyr Val Thr Trp Asn Lys Gly Glu Asp Ser Trp Gln Asp Met
              210 215 220
          Met Leu Met Ser His Cys Arg His His Ile Ile Cys Asn Ser Thr Phe
          225 230 235 240
          Ser Trp Trp Gly Ala Trp Leu Asn Pro Arg Lys Glu Lys Ile Val Ile
                          245 250 255
          Ala Pro Cys Arg Trp Phe Gln His Lys Glu Thr Pro Asp Met Tyr Pro
                      260 265 270
          Lys Glu Trp Ile Lys Val Pro Ile Asn
                  275 280
           <![CDATA[ <210> 104]]>
           <![CDATA[ <211> 290]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Akkermansia muciniphila]]>
           <![CDATA[ <400> 104]]>
          Met Arg Leu Phe Gly Gly Leu Gly Asn Gln Leu Phe Gln Tyr Ala Phe
          1 5 10 15
          Leu Phe Ala Leu Ser Arg Gln Gly Gly Lys Ala Arg Leu Glu Thr Ser
                      20 25 30
          Ser Tyr Glu His Asp Asp Lys Arg Val Cys Glu Leu His His Phe Arg
                  35 40 45
          Val Ser Leu Pro Ile Glu Gly Gly Pro Pro Pro Trp Ala Phe Arg Lys
              50 55 60
          Ser Arg Ile Pro Ala Cys Leu Arg Ser Leu Phe Ala Ala Pro Lys Tyr
          65 70 75 80
          Pro His Phe Arg Glu Glu Lys Arg His Gly Phe Asp Pro Gly Leu Ala
                          85 90 95
          Ala Pro Pro Arg Arg His Thr Tyr Phe Lys Gly Tyr Phe Gln Thr Glu
                      100 105 110
          Gln Tyr Phe Leu His Cys Arg Glu Gln Leu Cys Arg Glu Phe Arg Leu
                  115 120 125
          Lys Thr Pro Leu Thr Pro Glu Asn Ala Arg Ile Leu Glu Asp Ile Arg
              130 135 140
          Ser Cys Cys Ser Ile Ser Leu His Ile Arg Arg Thr Asp Tyr Leu Ser
          145 150 155 160
          Asn Pro Tyr Leu Ser Pro Pro Pro Leu Glu Tyr Tyr Leu Arg Ser Met
                          165 170 175
          Ala Glu Met Glu Gly Arg Leu Arg Ala Ala Gly Ala Pro Gln Glu Ser
                      180 185 190
          Leu Arg Tyr Phe Ile Phe Ser Asp Asp Ile Glu Trp Ala Arg Gln Asn
                  195 200 205
          Leu Arg Pro Ala Leu Pro His Val His Val Asp Ile Asn Asp Gly Gly
              210 215 220
          Thr Gly Tyr Phe Asp Leu Glu Leu Met Arg Asn Cys Arg His His Ile
          225 230 235 240
          Ile Ala Asn Ser Thr Phe Ser Trp Trp Ala Ala Trp Leu Asn Glu His
                          245 250 255
          Ala Glu Lys Ile Val Ile Ala Pro Arg Ile Trp Phe Asn Arg Glu Glu
                      260 265 270
          Gly Asp Arg Tyr His Thr Asp Asp Ala Leu Ile Pro Gly Ser Trp Leu
                  275 280 285
          Arg Ile
              290
           <![CDATA[ <210> 105]]>
           <![CDATA[ <211> 298]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Salmonella enterica]]>
           <![CDATA[ <400> 105]]>
          Met Tyr Ser Cys Leu Ser Gly Gly Leu Gly Asn Gln Met Phe Gln Tyr
          1 5 10 15
          Ala Ala Ala Tyr Ile Leu Lys Gln Tyr Phe Gln Ser Thr Thr Leu Val
                      20 25 30
          Leu Asp Asp Ser Tyr Tyr Tyr Ser Gln Pro Lys Arg Asp Thr Val Arg
                  35 40 45
          Ser Leu Glu Leu Asn Gln Phe Asn Ile Ser Tyr Asp Arg Phe Ser Phe
              50 55 60
          Ala Asp Glu Lys Glu Lys Ile Lys Leu Leu Arg Lys Phe Lys Arg Asn
          65 70 75 80
          Pro Phe Pro Lys Gln Ile Ser Glu Ile Leu Ser Ile Ala Leu Phe Gly
                          85 90 95
          Lys Tyr Ala Leu Ser Asp Arg Ala Phe Tyr Thr Phe Glu Thr Ile Lys
                      100 105 110
          Asn Ile Asp Lys Ala Cys Leu Phe Ser Phe Tyr Gln Asp Ala Asp Leu
                  115 120 125
          Leu Asn Lys Tyr Lys Gln Leu Ile Leu Pro Leu Phe Glu Leu Arg Asp
              130 135 140
          Asp Leu Leu Asp Ile Cys Lys Asn Leu Glu Leu Tyr Ser Leu Ile Gln
          145 150 155 160
          Arg Ser Asn Asn Thr Thr Ala Leu His Ile Arg Arg Gly Asp Tyr Val
                          165 170 175
          Thr Asn Gln His Ala Ala Lys Tyr His Gly Val Leu Asp Ile Ser Tyr
                      180 185 190
          Tyr Asn His Ala Met Glu Tyr Val Glu Arg Glu Arg Gly Lys Gln Asn
                  195 200 205
          Phe Ile Ile Phe Ser Asp Asp Val Arg Trp Ala Gln Lys Ala Phe Leu
              210 215 220
          Glu Asn Asp Asn Cys Tyr Val Ile Asn Asn Ser Asp Tyr Asp Phe Ser
          225 230 235 240
          Ala Ile Asp Met Tyr Leu Met Ser Leu Cys Lys Asn Asn Ile Ile Ala
                          245 250 255
          Asn Ser Thr Tyr Ser Trp Trp Gly Ala Trp Leu Asn Lys Tyr Glu Asp
                      260 265 270
          Lys Leu Val Ile Ser Pro Lys Gln Trp Phe Leu Gly Asn Asn Glu Thr
                  275 280 285
          Ser Leu Arg Asn Ala Ser Trp Ile Thr Leu
              290 295
           <![CDATA[ <210> 106]]>
           <![CDATA[ <211> 281]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Bacteroides sp. CAG:633]]>
           <![CDATA[ <400> 106]]>
          Met Arg Leu Ile Lys Met Thr Gly Gly Leu Gly Asn Gln Met Phe Ile
          1 5 10 15
          Tyr Ala Phe Tyr Leu Arg Met Lys Lys Arg Tyr Pro Lys Val Arg Ile
                      20 25 30
          Asp Leu Ser Asp Met Val His Tyr His Val His His Gly Tyr Glu Met
                  35 40 45
          His Arg Val Phe Asn Leu Pro His Thr Glu Phe Cys Ile Asn Gln Pro
              50 55 60
          Leu Lys Lys Val Ile Glu Phe Leu Phe Phe Lys Lys Ile Tyr Glu Arg
          65 70 75 80
          Lys Gln Asp Pro Asn Ser Leu Arg Ala Phe Glu Lys Lys Tyr Leu Trp
                          85 90 95
          Pro Leu Leu Tyr Phe Lys Gly Phe Tyr Gln Ser Glu Arg Phe Phe Ala
                      100 105 110
          Asp Ile Lys Asp Glu Val Arg Lys Ala Phe Thr Phe Asp Ser Ser Lys
                  115 120 125
          Val Asn Ala Arg Ser Ala Glu Leu Leu Arg Arg Leu Asp Ala Asp Ala
              130 135 140
          Asn Ala Val Ser Leu His Ile Arg Arg Gly Asp Tyr Leu Gln Pro Gln
          145 150 155 160
          His Trp Ala Thr Thr Gly Ser Val Cys Gln Leu Pro Tyr Tyr Gln Asn
                          165 170 175
          Ala Ile Ala Glu Met Asn Arg Arg Val Ala Ala Pro Ser Tyr Tyr Val
                      180 185 190
          Phe Ser Asp Asp Ile Ala Trp Val Lys Glu Asn Ile Pro Leu Gln Asn
                  195 200 205
          Ala Val Tyr Ile Asp Trp Asn Lys Gly Glu Glu Ser Trp Gln Asp Met
              210 215 220
          Met Leu Met Ser His Cys Arg His His Ile Ile Cys Asn Ser Thr Phe
          225 230 235 240
          Ser Trp Trp Gly Ala Trp Leu Asp Pro His Glu Asp Lys Ile Val Ile
                          245 250 255
          Val Pro Asn Arg Trp Phe Gln His Cys Glu Thr Pro Asn Ile Tyr Pro
                      260 265 270
          Ala Gly Trp Val Lys Val Ala Ile Asn
                  275 280
           <![CDATA[ <210> 107]]>
           <![CDATA[ <211> 291]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Bacteroides common]]>
           <![CDATA[ <400> 107]]>
          Met Gln Ile Thr Lys Lys Lys Gly Gln Arg Asn Met Arg Leu Ile Lys Val
          1 5 10 15
          Thr Gly Gly Leu Gly Asn Gln Met Phe Ile Tyr Ala Phe Tyr Leu Arg
                      20 25 30
          Met Lys Lys Tyr Tyr Pro Lys Val Arg Ile Asp Leu Ser Asp Met Met
                  35 40 45
          His Tyr Lys Val His Tyr Gly Tyr Glu Met His Arg Val Phe Asn Leu
              50 55 60
          Pro His Thr Glu Phe Cys Ile Asn Gln Pro Leu Lys Lys Val Ile Glu
          65 70 75 80
          Phe Leu Phe Phe Lys Lys Ile Tyr Glu Arg Lys Gln Ala Pro Asn Ser
                          85 90 95
          Leu Arg Ala Phe Glu Lys Lys Tyr Phe Trp Pro Leu Leu Tyr Phe Lys
                      100 105 110
          Gly Phe Tyr Gln Ser Glu Arg Phe Phe Ala Asp Ile Lys Asp Glu Val
                  115 120 125
          Arg Glu Ser Phe Thr Phe Asp Lys Asn Lys Ala Asn Ser Arg Ser Leu
              130 135 140
          Asn Met Leu Glu Ile Leu Asp Lys Asp Glu Asn Ala Val Ser Leu His
          145 150 155 160
          Ile Arg Arg Gly Asp Tyr Leu Gln Pro Lys His Trp Ala Thr Thr Gly
                          165 170 175
          Ser Val Cys Gln Leu Pro Tyr Tyr Gln Asn Ala Ile Ala Glu Met Ser
                      180 185 190
          Arg Arg Val Ala Ser Pro Ser Tyr Tyr Ile Phe Ser Asp Asp Ile Ala
                  195 200 205
          Trp Val Lys Glu Asn Leu Pro Leu Gln Asn Ala Val Tyr Ile Asp Trp
              210 215 220
          Asn Thr Asp Glu Asp Ser Trp Gln Asp Met Met Leu Met Ser His Cys
          225 230 235 240
          Lys His His Ile Ile Cys Asn Ser Thr Phe Ser Trp Trp Gly Ala Trp
                          245 250 255
          Leu Asn Pro Asn Met Asp Lys Thr Val Ile Val Pro Ser Arg Trp Phe
                      260 265 270
          Gln His Ser Glu Ala Pro Asp Ile Tyr Pro Thr Gly Trp Ile Lys Val
                  275 280 285
          Pro Val Ser
              290
           <![CDATA[ <210> 108]]>
           <![CDATA[ <211> 300]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Helicobacter pylori]]>
           <![CDATA[ <400> 108]]>
          Met Ala Phe Lys Val Val Gln Ile Cys Gly Gly Leu Gly Asn Gln Met
          1 5 10 15
          Phe Gln Tyr Ala Phe Ala Lys Ser Leu Gln Lys His Leu Asn Thr Pro
                      20 25 30
          Val Leu Leu Asp Thr Thr Ser Phe Asp Trp Ser Asn Arg Lys Met Gln
                  35 40 45
          Leu Glu Leu Phe Pro Ile Asp Leu Pro Tyr Ala Asn Ala Lys Glu Ile
              50 55 60
          Ala Ile Ala Lys Met Gln His Leu Pro Lys Leu Val Arg Asp Ala Leu
          65 70 75 80
          Lys Tyr Ile Gly Phe Asp Arg Val Ser Gln Glu Ile Val Phe Glu Tyr
                          85 90 95
          Glu Pro Lys Leu Leu Lys Pro Ser Arg Leu Thr Tyr Phe Phe Gly Tyr
                      100 105 110
          Phe Gln Asp Pro Arg Tyr Phe Asp Ala Ile Ser Ser Leu Ile Lys Gln
                  115 120 125
          Thr Phe Thr Leu Pro Pro Pro Pro Glu Asn Asn Lys Asn Asn Asn Lys
              130 135 140
          Lys Glu Glu Glu Tyr Gln Arg Lys Leu Ser Leu Ile Leu Ala Ala Lys
          145 150 155 160
          Asn Ser Val Phe Val His Ile Arg Arg Gly Asp Tyr Val Gly Ile Gly
                          165 170 175
          Cys Gln Leu Gly Ile Asp Tyr Gln Lys Lys Ala Leu Glu Tyr Met Ala
                      180 185 190
          Lys Arg Val Pro Asn Met Glu Leu Phe Val Phe Cys Glu Asp Leu Lys
                  195 200 205
          Phe Thr Gln Asn Leu Asp Leu Gly Tyr Pro Phe Thr Asp Met Thr Thr
              210 215 220
          Arg Asp Lys Glu Glu Glu Ala Tyr Trp Asp Met Leu Leu Met Gln Ser
          225 230 235 240
          Cys Lys His Gly Ile Ile Ala Asn Ser Thr Tyr Ser Trp Trp Ala Ala
                          245 250 255
          Tyr Leu Met Glu Asn Pro Glu Lys Ile Ile Ile Gly Pro Lys His Trp
                      260 265 270
          Leu Phe Gly His Glu Asn Ile Leu Cys Lys Glu Trp Val Lys Ile Glu
                  275 280 285
          Ser His Phe Glu Val Lys Ser Gln Lys Tyr Asn Ala
              290 295 300
           <![CDATA[ <210> 109]]>
           <![CDATA[ <211> 280]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Bacteroides sp. An51A]]>
           <![CDATA[ <400> 109]]>
          Met Arg Leu Ile Lys Val Thr Gly Gly Leu Gly Asn Gln Met Phe Ile
          1 5 10 15
          Tyr Ala Cys Tyr Leu Gln Met Lys Lys Arg Phe Pro Arg Val Tyr Ile
                      20 25 30
          Asp Leu Thr Asp Met Met His Tyr His Val His Tyr Gly Tyr Glu Met
                  35 40 45
          His Arg Val Phe Gly Leu Pro Cys Val Glu Ile Cys Met Asn Gln Val
              50 55 60
          Leu Lys Lys Val Ile Glu Phe Leu Phe Phe Lys Thr Ile Leu Glu Arg
          65 70 75 80
          Lys Gln His Gly Ser Met Arg Pro Tyr Thr Arg Ser Tyr Leu Trp Pro
                          85 90 95
          Leu Ile Tyr Phe Lys Gly Phe Tyr Gln Ser Glu Lys Tyr Phe Ala Asp
                      100 105 110
          Ile Lys Asp Glu Val Arg Gln Ala Phe Thr Phe Asp Leu Lys Gln Ala
                  115 120 125
          Asn Gly Gln Ser Leu Arg Met Arg Glu Gln Ile Asp Lys Asp Glu His
              130 135 140
          Ala Val Ser Leu His Val Arg Arg Gly Asp Tyr Leu Gln Pro Lys His
          145 150 155 160
          Trp Glu Ala Ile Gly Cys Ile Cys Gln Leu Ser Tyr Tyr Arg Asn Ala
                          165 170 175
          Leu Glu Glu Met Gly Lys Arg Val Glu Arg Pro Arg Tyr Tyr Val Phe
                      180 185 190
          Ser Asp Asp Leu Ala Trp Val Lys Glu Asn Leu Asp Leu Pro Asp Ala
                  195 200 205
          Val Tyr Ile Asp Trp Asn Lys Gly Glu Asp Ser Trp Gln Asp Met Met
              210 215 220
          Leu Met Ser Arg Cys Arg His His Ile Ile Cys Asn Ser Thr Phe Ser
          225 230 235 240
          Trp Trp Gly Ala Trp Leu Asn Pro Arg Asn Gly Lys Ile Val Ile Ala
                          245 250 255
          Pro Glu Arg Trp Thr Arg Asp Ala Asp Ser Arg Glu Ile Val Pro Ala
                      260 265 270
          Glu Trp Leu Arg Val Ser Ile Lys
                  275 280
           <![CDATA[ <210> 110]]>
           <![CDATA[ <211> 340]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Helicobacter sp. CLO-3]]>
           <![CDATA[ <400> 110]]>
          Met Thr Ile Ile Asn Ile Arg Asp Gly Leu Gly Asn Gln Met Phe Gln
          1 5 10 15
          Tyr Ala Phe Ala Lys Val Leu Glu Ser Lys Gly Glu Ser Val Val Leu
                      20 25 30
          Asp Thr Ser Trp Tyr Ser Glu Glu Pro Asp Ser Val Ser Lys Asn Ile
                  35 40 45
          Ala Asn Lys Lys Asn Ile Arg Asn Leu Glu Ile Thr Arg Phe Asp Ile
              50 55 60
          Lys Ile Val Pro Tyr Met Glu Ile Glu Thr Met Leu Gln Thr Gln Asp
          65 70 75 80
          Lys Phe Tyr His Phe Phe Ser Phe Ile His Lys Leu Ile Ala Tyr Ala
                          85 90 95
          Pro Lys Glu Cys Leu Arg Val Gly Leu Lys Ser Phe Leu Pro Lys Thr
                      100 105 110
          Arg Arg Phe His Pro Tyr Lys Tyr His Ile Lys Glu Val Tyr Asp Ser
                  115 120 125
          Arg Asp Leu Ser Ala Leu Pro His Asp Ile Leu Lys Asn Ala Leu Ile
              130 135 140
          Phe Gly Phe Phe Gln Lys Leu Ser Tyr Phe Lys His Ile Asp Ser Glu
          145 150 155 160
          Ile Arg Ala Asp Phe Thr Leu Cys Thr Pro Leu Ser Pro Ala Asn Glu
                          165 170 175
          Ala Met Lys Lys Arg Ile Leu Ser Thr Pro Asn Ala Ala Phe Ile His
                      180 185 190
          Ile Arg Arg Gly Asp Tyr Leu Asn Val Trp Gln Val Ile Lys Leu Gly
                  195 200 205
          Lys Ala Tyr Tyr Ala Ser Ala Ile Lys Glu Ile Leu Thr His Val Lys
              210 215 220
          Asn Pro Lys Phe Phe Ile Phe Ser Asn Asp Ile Glu Trp Cys Lys Asn
          225 230 235 240
          Asn Phe Ile Asn Leu Ile Asp Ser Ser Val Phe Ala Gly Lys Glu Tyr
                          245 250 255
          Glu Phe Val Glu Gln Asn Gly Glu Gly Asp Ala Ile Glu Glu Leu Glu
                      260 265 270
          Leu Met Arg Ser Cys Lys His Gly Ile Ile Ala Asn Ser Thr Phe Ser
                  275 280 285
          Trp Trp Ala Ala Tyr Leu Met Glu Asn Pro Lys Lys Thr Val Ile Ala
              290 295 300
          Pro Ser Lys Phe Phe Leu Ile Pro Pro Pro Gln Glu Pro Asn His Ile
          305 310 315 320
          Asp Asp Ile Leu Pro Glu Gly Trp Ile Lys Thr Asp Pro Thr Trp Gly
                          325 330 335
          Asn Val Glu Ser
                      340
           <![CDATA[ <210> 111]]>
           <![CDATA[ <211> 478]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Helicobacter pylori]]>
           <![CDATA[ <400> 111]]>
          Met Phe Gln Pro Leu Leu Asp Ala Tyr Val Glu Ser Ala Ser Ile Glu
          1 5 10 15
          Lys Met Ala Ser Lys Ser Pro Pro Pro Leu Lys Ile Ala Val Ala Asn
                      20 25 30
          Trp Trp Gly Asp Glu Glu Ile Lys Glu Phe Lys Asn Ser Val Leu Tyr
                  35 40 45
          Phe Ile Leu Ser Gln Arg Tyr Thr Ile Thr Leu His Gln Asn Pro Asn
              50 55 60
          Glu Phe Ser Asp Leu Val Phe Gly Asn Pro Leu Gly Ser Ala Arg Lys
          65 70 75 80
          Ile Leu Ser Tyr Gln Asn Ala Lys Arg Val Phe Tyr Thr Gly Glu Asn
                          85 90 95
          Glu Ser Pro Asn Phe Asn Leu Phe Asp Tyr Ala Ile Gly Phe Asp Glu
                      100 105 110
          Leu Asp Phe Asn Asp Arg Tyr Leu Arg Met Pro Leu Tyr Tyr Asp Arg
                  115 120 125
          Leu His His Lys Ala Glu Ser Val Asn Asp Thr Thr Ala Pro Tyr Lys
              130 135 140
          Leu Lys Asp Asn Ser Leu Tyr Ala Leu Lys Lys Pro Ser His Cys Phe
          145 150 155 160
          Lys Glu Lys His Pro Asn Leu Cys Ala Val Val Asn Asp Glu Ser Asp
                          165 170 175
          Pro Leu Lys Arg Gly Phe Ala Ser Phe Val Ala Ser Asn Pro Asn Ala
                      180 185 190
          Pro Ile Arg Asn Ala Phe Tyr Asp Ala Leu Asn Ser Ile Glu Pro Val
                  195 200 205
          Thr Gly Gly Gly Ser Val Arg Asn Thr Leu Gly Tyr Asn Val Lys Asn
              210 215 220
          Lys Asn Glu Phe Leu Ser Gln Tyr Lys Phe Asn Leu Cys Phe Glu Asn
          225 230 235 240
          Thr Gln Gly Tyr Gly Tyr Val Thr Glu Lys Ile Ile Asp Ala Tyr Phe
                          245 250 255
          Ser His Thr Ile Pro Ile Tyr Trp Gly Ser Pro Ser Val Ala Lys Asp
                      260 265 270
          Phe Asn Pro Lys Ser Phe Val Asn Val His Asp Phe Lys Asn Phe Asp
                  275 280 285
          Glu Ala Ile Asp Tyr Ile Lys Tyr Leu His Thr His Lys Asn Ala Tyr
              290 295 300
          Leu Asp Met Leu Tyr Glu Asn Pro Leu Asn Thr Leu Asp Gly Lys Ala
          305 310 315 320
          Tyr Phe Tyr Gln Asn Leu Ser Phe Lys Lys Ile Leu Ala Phe Phe Lys
                          325 330 335
          Thr Ile Leu Glu Asn Asp Thr Ile Tyr His Asp Asn Pro Phe Ile Phe
                      340 345 350
          Cys Arg Asp Leu Asn Glu Pro Leu Val Thr Ile Asp Asp Leu Arg Val
                  355 360 365
          Asn Tyr Asp Asp Leu Arg Val Asn Tyr Asp Asp Leu Arg Ile Asn Tyr
              370 375 380
          Asp Asp Leu Arg Val Asn Tyr Asp Asp Leu Arg Ile Asn Tyr Asp Asp
          385 390 395 400
          Leu Arg Val Asn Tyr Asp Asp Leu Arg Val Asn Tyr Asp Asp Leu Arg
                          405 410 415
          Ile Asn Tyr Asp Asp Leu Arg Val Asn Tyr Asp Asp Leu Arg Val Asn
                      420 425 430
          Tyr Glu Arg Leu Leu Ser Lys Ala Thr Pro Leu Leu Glu Leu Ser Gln
                  435 440 445
          Asn Thr Thr Ser Lys Ile Tyr Arg Lys Ala Tyr Gln Lys Ser Leu Pro
              450 455 460
          Leu Leu Arg Ala Ile Arg Arg Trp Val Lys Lys Leu Gly Leu
          465 470 475
           <![CDATA[ <210> 112]]>
           <![CDATA[ <211> 425]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Helicobacter pylori]]>
           <![CDATA[ <400> 112]]>
          Met Phe Gln Pro Leu Leu Asp Ala Phe Ile Glu Ser Ala Ser Ile Glu
          1 5 10 15
          Lys Met Ala Ser Lys Ser Pro Pro Pro Pro Leu Lys Ile Ala Val Ala
                      20 25 30
          Asn Trp Trp Gly Asp Glu Glu Ile Lys Glu Phe Lys Lys Ser Val Leu
                  35 40 45
          Tyr Phe Ile Leu Ser Gln Arg Tyr Ala Ile Thr Leu His Gln Asn Pro
              50 55 60
          Asn Glu Phe Ser Asp Leu Val Phe Ser Asn Pro Leu Gly Ala Ala Arg
          65 70 75 80
          Lys Ile Leu Ser Tyr Gln Asn Thr Lys Arg Val Phe Tyr Thr Gly Glu
                          85 90 95
          Asn Glu Ser Pro Asn Phe Asn Leu Phe Asp Tyr Ala Ile Gly Phe Asp
                      100 105 110
          Glu Leu Asp Phe Asn Asp Arg Tyr Leu Arg Met Pro Leu Tyr Tyr Ala
                  115 120 125
          His Leu His Tyr Lys Ala Glu Leu Val Asn Asp Thr Thr Ala Pro Tyr
              130 135 140
          Lys Leu Lys Asp Asn Ser Leu Tyr Ala Leu Lys Lys Pro Ser His His
          145 150 155 160
          Phe Lys Glu Asn His Pro Asn Leu Cys Ala Val Val Asn Asp Glu Ser
                          165 170 175
          Asp Leu Leu Lys Arg Gly Phe Ala Ser Phe Val Ala Ser Asn Ala Asn
                      180 185 190
          Ala Pro Met Arg Asn Ala Phe Tyr Asp Ala Leu Asn Ser Ile Glu Pro
                  195 200 205
          Val Thr Gly Gly Gly Ser Val Arg Asn Thr Leu Gly Tyr Lys Val Gly
              210 215 220
          Asn Lys Ser Glu Phe Leu Ser Gln Tyr Lys Phe Asn Leu Cys Phe Glu
          225 230 235 240
          Asn Ser Gln Gly Tyr Gly Tyr Val Thr Glu Lys Ile Leu Asp Ala Tyr
                          245 250 255
          Phe Ser His Thr Ile Pro Ile Tyr Trp Gly Ser Pro Ser Val Ala Lys
                      260 265 270
          Asp Phe Asn Pro Lys Ser Phe Val Asn Val His Asp Phe Asn Asn Phe
                  275 280 285
          Asp Glu Ala Ile Asp Tyr Ile Lys Tyr Leu His Thr His Pro Asn Ala
              290 295 300
          Tyr Leu Asp Met Leu Tyr Glu Asn Pro Leu Asn Thr Leu Asp Gly Lys
          305 310 315 320
          Ala Tyr Phe Tyr Gln Asp Leu Ser Phe Lys Lys Ile Leu Asp Phe Phe
                          325 330 335
          Lys Thr Ile Leu Glu Asn Asp Thr Ile Tyr His Lys Phe Ser Thr Ser
                      340 345 350
          Phe Met Trp Glu Tyr Asp Leu His Lys Pro Leu Val Ser Ile Asp Asp
                  355 360 365
          Leu Arg Val Asn Tyr Asp Asp Leu Arg Val Asn Tyr Asp Arg Leu Leu
              370 375 380
          Gln Asn Ala Ser Pro Leu Leu Glu Leu Ser Gln Asn Thr Thr Phe Lys
          385 390 395 400
          Ile Tyr Arg Lys Ala Tyr Gln Lys Ser Leu Pro Leu Leu Arg Ala Val
                          405 410 415
          Arg Lys Leu Val Lys Lys Leu Gly Leu
                      420 425
           <![CDATA[ <210> 113]]>
           <![CDATA[ <211> 329]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Basel psittaci pulmonary]]>
           <![CDATA[ <400> 113]]>
          Met Ser Val Leu Lys Lys Leu Val Arg Thr Leu Lys Lys Lys Lys Lys Asp
          1 5 10 15
          Ile Pro Ser Glu Asn Gln Glu Asp Ile Lys Pro Gln Glu Phe Gly His
                      20 25 30
          Ile Lys His Tyr His Phe Trp Pro Leu Ser Asn Glu Thr Phe Phe Asn
                  35 40 45
          Gln Phe Ala Gln Glu Lys Asn Leu Asp Leu Ser Gln Thr Ala Leu Ile
              50 55 60
          Ser Cys Phe Gly Glu Leu Ser Ala Ile Pro Lys Ile Pro Glu Arg Tyr
          65 70 75 80
          Lys Val Phe Phe Thr Gly Glu Asn Ile Tyr His Pro Asp Arg Ile Ser
                          85 90 95
          Tyr Ser Asp Pro Glu Leu Tyr Arg Met Val Asp Leu Tyr Leu Gly Phe
                      100 105 110
          Glu Tyr Arg Thr Glu Pro Lys Tyr Leu Arg Phe Pro Leu Trp Val Trp
                  115 120 125
          Tyr Leu Cys Gly Leu Thr Lys Lys Pro His Phe Ser His Glu Ser Ile
              130 135 140
          Ala Glu Phe Ile Arg Lys Met Asn Gln Pro Glu Phe Arg Leu Gln Ser
          145 150 155 160
          Ser Arg Asn Arg Phe Cys Ser His Ile Ser Ser His Asp Thr Asn Gly
                          165 170 175
          Ile Arg Lys Arg Met Ile Asp Leu Ile Leu Pro Ile Ala Ser Val Asp
                      180 185 190
          Cys Ala Gly Lys Phe Met Asn Asn Thr Asp Glu Leu Lys Ala Lys Phe
                  195 200 205
          Asn Asp Asp Lys Ile Asp Tyr Leu Lys Gln Tyr Arg Phe Asn Leu Cys
              210 215 220
          Pro Glu Asn Ser Glu Ser Val Gly Tyr Ile Thr Glu Lys Ile Phe Glu
          225 230 235 240
          Ser Ile Met Ala Gly Cys Ile Pro Ile Tyr Trp Gly Gly Val Lys Gln
                          245 250 255
          Leu Phe Val Glu Pro Asp Ile Leu Asn Pro Glu Ala Phe Ile Tyr Tyr
                      260 265 270
          Glu Lys Gly Lys Glu Glu Gln Leu Ala Lys Gln Val Glu Glu Leu Trp
                  275 280 285
          Ile Ser Pro Lys Arg Tyr Glu Glu Phe Ala Ala Ile Ala Pro Phe Lys
              290 295 300
          Glu Asp Ala Ala Glu Val Ile Tyr Thr Trp Ile Glu Glu Leu Glu Lys
          305 310 315 320
          Arg Leu Arg Ala Phe Glu Pro Lys Ala
                          325
           <![CDATA[ <210> 114]]>
           <![CDATA[ <211> 325]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Azospirillum oryzae A2P]]>
           <![CDATA[ <400> 114]]>
          Met Ile Asp Gln Arg Thr Ser Asp Phe Leu Ser Glu Phe Leu Ala Ser
          1 5 10 15
          Ser His Arg Asp Pro Ala Arg Leu Asp Ser Phe Leu Leu His Gly Pro
                      20 25 30
          Gly Arg Gly Ala Arg Ala Ala Lys Pro Arg Leu Lys Ile Ala Phe Phe
                  35 40 45
          Asp Phe Trp Pro Glu Phe Asp Pro Ala Ala Asn Phe Phe Val Asp Ile
              50 55 60
          Leu Ser Ala Arg Phe Asp Val Ser Val Val Asp Asn Asp Ser Asp Leu
          65 70 75 80
          Ala Ile Val Ser Val Phe Gly Thr Arg His Arg Glu Ala Arg Thr Ala
                          85 90 95
          Arg Ser Met Phe Phe Thr Gly Glu Asn Val Arg Pro Pro Leu Asp Gly
                      100 105 110
          Val Asp Met Ser Val Ser Phe Asp Arg Ile Asp Asp Pro Arg His Tyr
                  115 120 125
          Arg Leu Pro Leu Tyr Val Met His Ala Trp Asp His Arg Arg Glu Gly
              130 135 140
          Ala Thr Pro His Phe Cys Gln Ser Val Leu Pro Pro Val Pro Pro Thr
          145 150 155 160
          Arg Glu Glu Ala Ala Lys Arg Lys Phe Cys Ala Phe Leu Tyr Lys Asn
                          165 170 175
          Pro Asn Cys Ala Arg Arg Asn Asp Phe Phe Gln Met Leu Cys Ala Arg
                      180 185 190
          Arg His Val Glu Ser Val Gly Trp Leu Leu Asn Asn Thr Gly Ser Val
                  195 200 205
          Val Lys Met Gly Trp Leu Pro Lys Ile Arg Val Phe Ser Arg Tyr Arg
              210 215 220
          Phe Ala Phe Ala Phe Glu Asn Ala Ser His Pro Gly Tyr Leu Thr Glu
          225 230 235 240
          Lys Ile Leu Asp Ala Phe Gln Ala Gly Ala Val Pro Leu Tyr Trp Gly
                          245 250 255
          Asp Pro Gly Val Leu Arg Asp Val Ala Ala Gly Ser Phe Ile Asp Val
                      260 265 270
          Ser Arg Tyr Ala Ser Asp Glu Glu Ala Cys Asp Ala Ile Leu Ala Ala
                  275 280 285
          Asp Asp Asp Tyr Asp Thr Tyr Arg Arg Tyr Arg Ser Thr Pro Pro Phe
              290 295 300
          Leu Gly Ala Glu Asp Phe Tyr Phe Asp Ala Phe Arg Leu Ala Glu Trp
          305 310 315 320
          Ile Glu Ser Arg Leu
                          325
           <![CDATA[ <210> 115]]>
           <![CDATA[ <211> 342]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Pyrromonas catarrhalis]]>
           <![CDATA[ <400> 115]]>
          Met Pro Ile Tyr Asp Ile Lys Ala Met Asn Thr Pro Ser Lys Gln Pro
          1 5 10 15
          Leu Arg Glu Arg Leu His Met Met Arg Arg Arg Asn Arg Val Arg Lys
                      20 25 30
          Arg Ser Val Ile Ala Leu Ile Lys Ser His Leu Asp Ser Ser Arg Tyr
                  35 40 45
          Gln Asp Tyr Asn Trp Trp Asp Ser His Ala Ser Thr Phe Trp Leu Pro
              50 55 60
          Arg Phe Ile Asp Leu His Leu Glu Pro Lys Lys Lys Ile Asn Leu Phe
          65 70 75 80
          Ser Cys Phe Gln Asn Pro Leu Met Leu Ile Arg Tyr Tyr Lys Gly Val
                          85 90 95
          Lys Ile Phe Leu Ser Gly Glu Asn Leu Thr Asn Asn Glu His Phe Gly
                      100 105 110
          Phe His Pro Arg Met Leu Asp His Arg Ile Asn Glu Val Asp Leu Ala
                  115 120 125
          Leu Gly Phe Glu Phe Arg Lys Asp Pro Lys Tyr Tyr Arg Phe Pro Leu
              130 135 140
          Trp Ile Tyr Gln Asn Glu Phe Ile Ser Pro Ser Ala Ser Leu Glu Asp
          145 150 155 160
          Ile Cys Val Leu Val Gly Gln Ile Asn Asp Pro Ser Thr Arg Arg Ser
                          165 170 175
          Ala Lys Arg Ser Arg Phe Ile Gly Gln Ile Ser Ser His Asp Lys Gly
                      180 185 190
          Gly Met Arg Gly Arg Leu Ile Asp Leu Leu Ser Pro Ile Gly Gln Ile
                  195 200 205
          Asp Cys Ala Gly Lys Phe Arg His Asn Thr Asp Glu Leu Leu Glu Val
              210 215 220
          Tyr Gly Asp Asp Lys Phe Lys Tyr Leu Ala Asn Tyr Arg Phe Asn Leu
          225 230 235 240
          Cys Pro Glu Asn Ser Leu Gly Glu Gly Tyr Ile Thr Glu Lys Val Phe
                          245 250 255
          Asp Ser Ile Arg Ala Gly Cys Ile Pro Ile Tyr Trp Gly Ala Tyr Leu
                      260 265 270
          Glu Pro Gly Ile Leu Asn Pro Lys Ala Ile Leu Arg Phe Glu Glu Gly
                  275 280 285
          Lys Glu Gln Glu Phe Tyr Asn Arg Val Lys Glu Leu Trp Glu Asn Glu
              290 295 300
          Glu Ala Tyr Glu Gln Phe Ile Leu Glu Pro Pro Phe Val Glu Gly Ala
          305 310 315 320
          Ala Glu Arg Ile Trp Glu Ile Leu Gln Gly Leu Arg Glu Arg Leu Ala
                          325 330 335
          Pro Leu Val Glu Glu Gly
                      340
           <![CDATA[ <210> 116]]>
           <![CDATA[ <211> 323]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Pyrromonas sp.]]>
           <![CDATA[ <400> 116]]>
          Met Asn Ala Val Glu Arg Val Arg Asn Ile Leu Asn Tyr Cys Ile Asn
          1 5 10 15
          Glu Val Gln Met Tyr Arg Gln Cys Pro Asn Ser Lys Tyr Tyr Asn Phe
                      20 25 30
          Trp Pro Cys Asp Tyr Asn Asn Asn Trp Phe Asn His Phe Val Glu His
                  35 40 45
          Arg Gly Leu Ala Lys Glu Arg His Arg Leu Asn Phe Phe Ser Val Phe
              50 55 60
          Gly Asn Pro Leu Leu Pro Arg Ile Ile Pro Gly Lys Lys Val Phe Phe
          65 70 75 80
          Thr Gly Glu Asn Leu Ala Asp Asn Ser Ile His Ser Ile Gly Arg Ala
                          85 90 95
          Phe Lys Lys Thr Phe Pro Val Tyr Asp Leu Val Leu Gly Phe Asp Tyr
                      100 105 110
          Glu Val Glu Asp Ser Arg Val Asn Tyr Met Arg Phe Pro Leu Trp Ile
                  115 120 125
          Ala Phe Leu Ile Asp Pro Thr Ala Asp Tyr Gln Lys Ile Lys Glu Thr
              130 135 140
          Ile Glu Arg Ile Asn Asp Pro Ser Thr Arg Leu Asn Ala Ser Arg Asp
          145 150 155 160
          Arg Phe Ala Cys Leu Val Ala Ser His Asp Lys Thr Gly Ile Arg Gln
                          165 170 175
          Lys Leu Tyr Asp Val Leu Met Pro Ile Ala Ser Val Thr Cys Pro Gly
                      180 185 190
          Arg Phe Gln Asn Asn Thr Asn Glu Leu His Asp Leu Tyr Ala Asn Asp
                  195 200 205
          Lys Arg Glu Tyr Leu Lys Leu Phe Lys Phe Asn Val Cys Pro Glu Asn
              210 215 220
          Ser Ser Thr Pro Gly Tyr Ile Thr Glu Lys Leu Phe Asp Ser Phe Ala
          225 230 235 240
          Ser Gly Cys Ile Pro Ile Tyr Phe Gly Gly Gly Thr Glu Glu Ile Glu
                          245 250 255
          Pro Asp Ile Val Asn Gln Gly Ala Phe Ile Arg Tyr Trp Asp Asp Gly
                      260 265 270
          Arg Met Asp Trp Met Asp Thr Val Arg Glu Leu Trp Glu Ser Pro Ser
                  275 280 285
          Ala Tyr Arg Ala Val Ala Glu Ile Pro Phe Lys Glu Gln Ala Ala
              290 295 300
          Asp Val Ile Tyr Ala Tyr Met Glu Asn Leu His Asp Lys Leu Ala Ala
          305 310 315 320
          Ile Val Arg
           <![CDATA[ <210> 117]]>
           <![CDATA[ <211> 316]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Injured Lunamonas]]>
           <![CDATA[ <400> 117]]>
          Met Leu Met Arg Ala Leu Arg Lys Met Lys Arg Trp Gly Arg Val Ala
          1 5 10 15
          Phe Asp Tyr Thr Asn Thr Thr Lys Asp Gly Ala Val Cys Tyr His Asn
                      20 25 30
          Trp Trp Pro Cys Asn Tyr Glu Glu Glu Trp Phe His Arg Phe Val Val
                  35 40 45
          Gln Asn Ile Gly Thr Glu Arg Cys Tyr His Phe Phe Ser Val Phe Gly
              50 55 60
          Pro Arg Ile Ala Leu Thr Leu Pro Thr Pro Asn Lys Val Phe Phe Cys
          65 70 75 80
          Gly Glu Asn Val His Asn Ala Glu Trp Pro Tyr Lys Ser Tyr Gln Asp
                          85 90 95
          His Ala Leu Gly Asp Val Lys Leu Ala Leu Gly Tyr Asp Asp Ile Gln
                      100 105 110
          Asp Glu Arg Tyr Ile Arg Phe Pro Leu Trp Leu Leu Tyr Met Phe Asp
                  115 120 125
          Pro Val Val Asp Arg Tyr Ala Ile Arg Glu Arg Ile Glu Glu Ile Asn
              130 135 140
          His Ala Glu Asn Thr Arg Lys Tyr Glu Cys Val Leu Ile Ser Arg His
          145 150 155 160
          Asp Lys Trp Asn Met Arg Gly Pro Ile Tyr Asp Ala Leu Lys Asp His
                          165 170 175
          Leu Ala Ile Ser Cys Ala Gly Lys Trp Lys Gln Asn Thr Asp Glu Leu
                      180 185 190
          Trp Thr Val Tyr Asn Asp Asp Lys Pro Arg Tyr Leu Lys Glu Phe Lys
                  195 200 205
          Phe Asn Ile Cys Pro Glu Asn Phe Asp Thr Pro Tyr Tyr Val Thr Glu
              210 215 220
          Lys Leu Phe Glu Ala Phe Arg Ser Gly Thr Ile Pro Ile Tyr Ala Gly
          225 230 235 240
          Gly Gly Asp His Pro Glu Pro Glu Ile Val Asn Arg Ser Ala Leu Leu
                          245 250 255
          Leu Trp Glu Arg Gly Gln Ser Asp His Ser Ala Leu Val Gln Glu Val
                      260 265 270
          Ile Arg Leu Ala Arg Asp Glu Ile Tyr Tyr Asp Lys Phe Val His Gln
                  275 280 285
          Val Arg Leu Leu Pro Tyr Thr Glu Glu Phe Val Tyr Glu Gln Phe Ser
              290 295 300
          Ser Leu Lys Glu Arg Leu Leu Gln Ile Arg Arg Gly
          305 310 315
           <![CDATA[ <210> 118]]>
           <![CDATA[ <211> 325]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Adipogenic Azospirillum B510]]>
           <![CDATA[ <400> 118]]>
          Met Ile Asp Gln Arg Thr Ser Asp Phe Leu Ser Glu Phe Leu Ala Ser
          1 5 10 15
          Pro Asn Arg Asp Pro Ala Val Leu Asp Arg Phe Leu Leu His Gly Pro
                      20 25 30
          Glu Arg Gly Gly Arg Ala Ala Arg Pro Arg Leu Lys Ile Ala Phe Phe
                  35 40 45
          Asp Phe Trp Pro Glu Phe Asp Pro Ser Ala Asn Phe Phe Val Glu Ile
              50 55 60
          Leu Ser Ser Arg Phe Asp Val Ser Val Val Asp Asn Asp Ser Asp Leu
          65 70 75 80
          Ala Ile Leu Ser Val Phe Gly Glu Arg His Arg Glu Ala Arg Thr Ala
                          85 90 95
          Arg Ala Leu Phe Phe Thr Gly Glu Asn Val Arg Pro Pro Leu Asp Gly
                      100 105 110
          Val Asp Met Ser Val Ser Phe Asp Arg Ile Asp His Pro Arg His Tyr
                  115 120 125
          Arg Leu Pro Leu Tyr Val Met His Ala Trp Asp His Arg Arg Glu Gly
              130 135 140
          Ala Thr Pro His Phe Cys His Pro Val Leu Pro Pro Val Pro Pro Thr
          145 150 155 160
          Arg Glu Glu Ala Ala Lys Arg Lys Phe Cys Ala Phe Leu Tyr Lys Asn
                          165 170 175
          Pro His Cys Ala Arg Arg Asn Asp Phe Phe Gln Met Leu Cys Ala Arg
                      180 185 190
          Arg His Val Glu Ser Val Gly Trp Leu Leu Asn Asn Thr Gly Ser Val
                  195 200 205
          Val Lys Met Gly Trp Leu Pro Lys Ile Arg Val Phe Ala Arg Tyr Arg
              210 215 220
          Phe Ala Phe Ala Phe Glu Asn Ala Ala His Pro Gly Tyr Leu Thr Glu
          225 230 235 240
          Lys Ile Leu Asp Ala Phe Gln Ala Gly Thr Val Pro Leu Tyr Trp Gly
                          245 250 255
          Asp Ser Gly Val Leu Arg Asp Val Ala Ala Gly Ser Phe Ile Asp Val
                      260 265 270
          Ser Arg Tyr Ala Ser Asp Glu Glu Ala Ile Glu Ala Ile Leu Ala Ile
                  275 280 285
          Asp Asp Asp Tyr Asp Ser Tyr Arg Arg Tyr Arg Gly Thr Ala Pro Phe
              290 295 300
          Leu Gly Thr Glu Asp Phe Tyr Phe Asp Ala Tyr Arg Leu Ala Glu Trp
          305 310 315 320
          Ile Glu Ser Arg Leu
                          325
           <![CDATA[ <210> 119]]>
           <![CDATA[ <211> 325]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Azotobacter sp. TSH64]]>
           <![CDATA[ <400> 119]]>
          Met Ile Asp Gln Arg Thr Gly Val Phe Leu Ser Glu Phe Leu Asp Thr
          1 5 10 15
          Arg Asn Arg Asp Pro Ala Val Leu Asp Arg Phe Leu Leu Gln Gly Pro
                      20 25 30
          Asp Gly Gly Arg Arg Gly Ala Lys Pro Asn Leu Lys Val Ala Phe Phe
                  35 40 45
          Asp Phe Trp Pro Glu Phe Asp Pro Ser Ala Asn Phe Phe Val Glu Ile
              50 55 60
          Leu Ser Ala Arg Phe Gln Val Ser Val Val Glu Asn Asp Ser Asp Leu
          65 70 75 80
          Ala Ile Val Ser Val Phe Gly Thr Gly Pro Arg Glu Ile Arg Thr Ala
                          85 90 95
          Arg Ser Met Phe Phe Thr Gly Glu Asn Val Arg Pro Pro Leu Asp Gly
                      100 105 110
          Ile Asp Met Ser Val Ser Phe Asp Arg Ile Asp Asp Pro Arg His Phe
                  115 120 125
          Arg Leu Pro Leu Tyr Val Val His Ala Tyr Asp His Leu Arg Glu Gly
              130 135 140
          Ala Ala Pro Tyr Phe Cys Gln Pro Val Leu Pro Pro Val Pro Pro Thr
          145 150 155 160
          Arg Glu Asp Ala Ala Glu Arg Lys Phe Cys Ala Phe Leu Tyr Lys Asn
                          165 170 175
          Pro Asn Cys Ala Arg Arg Asn Asp Phe Phe His Met Leu Gly Ala Arg
                      180 185 190
          Arg His Val Asp Ser Val Gly Trp Leu Leu Asn Asn Thr Gly Ser Val
                  195 200 205
          Val Lys Met Gly Trp Leu Pro Lys Ile Arg Val Phe Ser Arg Tyr Arg
              210 215 220
          Phe Ala Phe Ala Phe Glu Asn Ala Ser His Pro Gly Tyr Leu Thr Glu
          225 230 235 240
          Lys Ile Leu Asp Ala Phe Gln Ala Gly Ala Val Pro Leu Tyr Trp Gly
                          245 250 255
          Asp Pro Gly Val Leu Arg Asp Val Ala Ala Gly Ser Phe Ile Asp Val
                      260 265 270
          Ser Arg Tyr Ser Ser Asp Glu Glu Ala Ile Glu Ala Ile Leu Ala Ile
                  275 280 285
          Asp Asp Asp Tyr Gly Ala Tyr Arg Arg Tyr Arg Ser Thr Pro Pro Phe
              290 295 300
          Leu Gly Thr Glu Asp Phe His Phe Asp Ala Tyr Arg Leu Ala Glu Trp
          305 310 315 320
          Ile Glu Ser Arg Leu
                          325
           <![CDATA[ <210> 120]]>
           <![CDATA[ <211> 325]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Azotobacter sp. TSH100]]>
           <![CDATA[ <400> 120]]>
          Met Ile Asp Arg Arg Thr Ser Asp Phe Leu Ala Glu Phe Leu Ala Ser
          1 5 10 15
          Ala Asn Lys Asp Pro Ala Val Leu Asp Arg Phe Leu Leu His Gly Pro
                      20 25 30
          Asp Arg Gly Gly Arg Ser Ala Lys Pro Arg Leu Lys Ile Ala Phe Phe
                  35 40 45
          Asp Phe Trp Pro Glu Phe Asp Pro Ala Ala Asn Phe Phe Val Glu Ile
              50 55 60
          Leu Ser Ala Arg Phe Asp Leu Ser Val Val Asp Asn Asp Ser Asp Leu
          65 70 75 80
          Ala Ile Val Ser Val Phe Gly Ile Arg His Arg Glu Ala Arg Thr Ala
                          85 90 95
          Arg Ser Leu Phe Phe Thr Gly Glu Asn Val Arg Pro Pro Leu Asp Gly
                      100 105 110
          Val Asp Met Ser Val Ser Phe Asp Arg Ile Asp Asp Pro Arg His Tyr
                  115 120 125
          Arg Leu Pro Leu Tyr Val Met His Ala Trp Asp His Arg Arg Glu Gly
              130 135 140
          Ala Thr Arg His Phe Cys His Ser Val Leu Pro Pro Val Pro Pro Thr
          145 150 155 160
          Arg Glu Glu Ala Asp Arg Arg Lys Phe Cys Ala Phe Leu Tyr Lys Asn
                          165 170 175
          Pro Asn Cys Glu Arg Arg Asn Asp Phe Phe Arg Met Leu Cys Ala Arg
                      180 185 190
          Arg His Val Glu Ser Val Gly Trp Leu Leu Asn Asn Thr Gly Ser Val
                  195 200 205
          Val Lys Met Gly Trp Leu Pro Lys Ile Arg Val Phe Ser Arg Tyr Arg
              210 215 220
          Phe Ala Phe Ala Phe Glu Asn Ala Ser His Pro Gly Tyr Leu Thr Glu
          225 230 235 240
          Lys Ile Leu Asp Ala Phe Gln Ala Gly Ala Val Pro Leu Tyr Trp Gly
                          245 250 255
          Asp Pro Gly Val Leu Arg Asp Val Ala Ala Gly Ser Phe Ile Asp Val
                      260 265 270
          Ser Arg Tyr Ser Ser Asp Glu Glu Ala Ile Asp Ala Ile Leu Ala Ile
                  275 280 285
          Asp Asp Asp Tyr Asp Thr Tyr Arg Arg His Arg Ser Thr Ala Pro Phe
              290 295 300
          Leu Gly Thr Glu Asp Phe Tyr Phe Asp Ala Phe Arg Leu Ala Glu Trp
          305 310 315 320
          Ile Glu Ser Arg Leu
                          325
           <![CDATA[ <210> 121]]>
           <![CDATA[ <211> 325]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Azospirillum brasiliensis]]>
           <![CDATA[ <400> 121]]>
          Met Leu Asp Gln Arg Thr Ser Ala Phe Leu Glu Glu Phe Leu Ala Lys
          1 5 10 15
          Pro Gly Gly Asp Pro Glu Arg Leu Asp Arg Phe Leu Leu His Gly Pro
                      20 25 30
          Tyr Arg Gly Arg Arg Gly Gly Arg Pro Arg Leu Lys Leu Ala Phe Tyr
                  35 40 45
          Asp Phe Trp Pro Glu Phe Asp Thr Gly Arg Asn Phe Phe Ile Glu Ile
              50 55 60
          Leu Ser Ser Arg Phe Asp Leu Ser Val Val Glu Asp Asp Ser Asp Leu
          65 70 75 80
          Ala Ile Val Ser Val Phe Gly Gly Arg His Arg Ala Ala Arg Ser Arg
                          85 90 95
          Arg Thr Leu Phe Phe Thr Gly Glu Asn Val Arg Pro Pro Leu Asp Gly
                      100 105 110
          Phe Asp Met Ala Val Ser Phe Asp Arg Val Gly Asp Pro Arg His Tyr
                  115 120 125
          Arg Leu Pro Leu Tyr Val Met His Ala Tyr Glu His Met Arg Glu Gly
              130 135 140
          Ala Val Pro His Phe Cys Ser Pro Val Leu Pro Pro Val Pro Pro Ser
          145 150 155 160
          Arg Ala Ala Phe Ala Glu Arg Asn Phe Cys Ala Phe Leu Tyr Lys Asn
                          165 170 175
          Pro Asn Gly Glu Arg Arg Asn Arg Phe Phe Pro Ala Leu Asp Ala Arg
                      180 185 190
          Arg Arg Val Asp Ser Val Gly Trp His Leu Asn Asn Thr Gly Ser Val
                  195 200 205
          Val Lys Met Gly Trp Leu Ala Lys Ile Arg Val Phe Glu Arg Tyr Arg
              210 215 220
          Phe Ala Phe Ala Phe Glu Asn Ala Ser His Pro Gly Tyr Leu Thr Glu
          225 230 235 240
          Lys Ile Leu Asp Val Phe Gln Ala Gly Ala Val Pro Leu Tyr Trp Gly
                          245 250 255
          Asp Pro Asp Val Glu Arg Glu Val Ala Ala Gly Ser Phe Ile Asp Val
                      260 265 270
          Ser Arg Phe Ala Thr Asp Glu Glu Ala Ala Glu His Ile Leu Ala Leu
                  275 280 285
          Asp Gly Asp Tyr Asp Ala Tyr Cys Ala Tyr Arg Ala Val Ala Pro Phe
              290 295 300
          Leu Gly Thr Glu Glu Phe His Phe Asp Ala Tyr Arg Leu Ala Asp Trp
          305 310 315 320
          Ile Glu Ser Arg Leu
                          325
           <![CDATA[ <210> 122]]>
           <![CDATA[ <211> 303]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Azospira sp. B510]]>
           <![CDATA[ <400> 122]]>
          Met Leu Asp Arg Phe Leu Leu His Gly Pro Glu Arg Gly Gly Arg Ala
          1 5 10 15
          Ala Arg Pro Arg Leu Lys Ile Ala Phe Phe Asp Phe Trp Pro Glu Phe
                      20 25 30
          Asp Pro Ser Ala Asn Phe Phe Val Glu Ile Leu Ser Ser Arg Phe Asp
                  35 40 45
          Val Ser Val Val Asp Asn Asp Ser Asp Leu Ala Ile Leu Ser Val Phe
              50 55 60
          Gly Glu Arg His Arg Glu Ala Arg Thr Ala Arg Ala Leu Phe Phe Thr
          65 70 75 80
          Gly Glu Asn Val Arg Pro Pro Leu Asp Gly Val Asp Met Ser Val Ser
                          85 90 95
          Phe Asp Arg Ile Asp His Pro Arg His Tyr Arg Leu Pro Leu Tyr Val
                      100 105 110
          Met His Ala Trp Asp His Arg Arg Glu Gly Ala Thr Pro His Phe Cys
                  115 120 125
          His Pro Val Leu Pro Pro Val Pro Pro Thr Arg Glu Glu Ala Ala Lys
              130 135 140
          Arg Lys Phe Cys Ala Phe Leu Tyr Lys Asn Pro His Cys Ala Arg Arg
          145 150 155 160
          Asn Asp Phe Phe Gln Met Leu Cys Ala Arg Arg His Val Glu Ser Val
                          165 170 175
          Gly Trp Leu Leu Asn Asn Thr Gly Ser Val Val Lys Met Gly Trp Leu
                      180 185 190
          Pro Lys Ile Arg Val Phe Ala Arg Tyr Arg Phe Ala Phe Ala Phe Glu
                  195 200 205
          Asn Ala Ala His Pro Gly Tyr Leu Thr Glu Lys Ile Leu Asp Ala Phe
              210 215 220
          Gln Ala Gly Thr Val Pro Leu Tyr Trp Gly Asp Ser Gly Val Leu Arg
          225 230 235 240
          Asp Val Ala Ala Gly Ser Phe Ile Asp Val Ser Arg Tyr Ala Ser Asp
                          245 250 255
          Glu Glu Ala Ile Glu Ala Ile Leu Ala Ile Asp Asp Asp Tyr Asp Ser
                      260 265 270
          Tyr Arg Arg Tyr Arg Gly Thr Ala Pro Phe Leu Gly Thr Glu Asp Phe
                  275 280 285
          Tyr Phe Asp Ala Tyr Arg Leu Ala Glu Trp Ile Glu Ser Arg Leu
              290 295 300
           <![CDATA[ <210> 123]]>
           <![CDATA[ <211> 309]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Shilong tail plank mold]]>
           <![CDATA[ <400> 123]]>
          Met Gln Lys Thr Phe Ser Leu Arg Pro Val Pro Val Asp Ala Ile Arg
          1 5 10 15
          Trp Asn Phe Ala Asn Phe Trp Ser Gly Phe Asp Ala Leu Ala Phe Glu
                      20 25 30
          Arg His Leu Leu Gly Val Ser Gly Lys His Lys Phe Arg Ile Ser Glu
                  35 40 45
          Gln Asn Pro Gln Ile Val Phe Glu Ser Val Phe Gly Thr Pro Gly Lys
              50 55 60
          Gly Arg Glu Arg Trp Pro Lys Ala Arg Gln Val Trp Tyr Thr Gly Glu
          65 70 75 80
          Asn Val Ala Pro Pro Leu Asp Gln Phe Asp Lys Cys Leu Ser Phe His
                          85 90 95
          Arg Asp Ile Lys Asp Pro Arg His Leu Arg Trp Pro Tyr Tyr Leu Leu
                      100 105 110
          His Leu Ala Ser Leu Pro Met Ser Phe Asn Asp Leu Val Lys Cys Gln
                  115 120 125
          Ser Ser Val Ser Thr Trp Ala Glu Arg Pro Gly Phe Cys Ala Phe Ile
              130 135 140
          Ala Phe Asn Glu Gly Cys Gln Thr Arg Asn Arg Phe Val Glu Lys Leu
          145 150 155 160
          Ser Arg Tyr Arg Arg Val Asp Cys Pro Gly Arg Val Leu Asn Asn Met
                          165 170 175
          Thr Ser Glu Thr Leu Gly Gln Arg Gly Asn Leu His Gly Lys Ile Asn
                      180 185 190
          Phe Leu Lys Gln Tyr Lys Tyr Ala Val Cys Phe Glu Asn Thr Ser Thr
                  195 200 205
          Arg Gly Ser Glu Gly Tyr Val Thr Glu Lys Leu Val Asp Ala Met Leu
              210 215 220
          Ala Gly Cys Ile Pro Leu Tyr Trp Gly Asp His Arg Val Gly Glu Asp
          225 230 235 240
          Phe Asn Glu Asn Ser Phe Ile Asn Leu Gly Val Tyr Gly Asn Asp Val
                          245 250 255
          Asn Ala Met Val Gln His Val Ile Glu Leu Asp Ser Asp Glu Arg Leu
                      260 265 270
          Gln Asn Asn Leu Phe Gln Glu Pro Trp Leu Pro Glu Ile Lys Ser Ser
                  275 280 285
          Glu His Phe Ser Phe Glu Thr Ser Lys Asp Ala Ile Leu Lys Leu Val
              290 295 300
          Ala Asn Val Asn Lys
          305
           <![CDATA[ <210> 124]]>
           <![CDATA[ <211> 316]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Geobacillus glucoside lysolytica]]>
           <![CDATA[ <400> 124]]>
          Met Lys Tyr Phe Leu Leu Leu Ala Lys Arg His Lys Lys Tyr Leu Lys
          1 5 10 15
          Glu Lys Lys Ile Phe Arg Asn Ser Thr Ile Ser Phe Tyr Asn Phe Trp
                      20 25 30
          Glu Ile Glu Asp Tyr Asn Asn Phe Trp Leu Gln Lys Phe Ile Val Asp
                  35 40 45
          Arg Asn Leu Asn Pro Lys Asn Lys Ser Ile Asn Phe Phe Ser Val Phe
              50 55 60
          Gly Pro Arg Tyr Val Leu Lys Lys Gln Lys Ala Ala Ile Asn Ile Phe
          65 70 75 80
          Phe Ser Gly Glu Thr Met Ser Arg Phe Lys Lys Tyr His Asp Tyr Cys
                          85 90 95
          Leu Pro Glu Val Asp Leu Ala Leu Gly Phe Asp Asp Leu Gln His Glu
                      100 105 110
          Lys Tyr Phe Arg Leu Pro Leu Trp Ile Leu Asp Phe Phe Glu Pro Thr
                  115 120 125
          Val Asp Leu Glu Lys Ala Lys Glu Lys Leu Lys Gln Leu Asn Tyr Tyr
              130 135 140
          Lys Asn Asn Lys Pro Ile Val Arg Glu Lys Phe Cys Ser Leu Ile Ala
          145 150 155 160
          Arg His Asp Glu Asn Gly Ile Arg Lys Lys Ile Val Asn Thr Leu Asn
                          165 170 175
          Pro Ile Glu Thr Val Asp Cys Ala Gly Lys Leu Phe Asn Asn Thr Ala
                      180 185 190
          Arg Leu Gln Thr Glu Phe Ala Asn Asn Lys Val Lys Phe Leu Glu Asn
                  195 200 205
          Tyr Lys Phe Asn Ile Cys Pro Glu Asn Thr Asn Gln Glu Ser Tyr Thr
              210 215 220
          Thr Glu Lys Leu Phe Glu Ser Phe Ala Ala Gly Cys Ile Pro Ile Tyr
          225 230 235 240
          Trp Gly Ser Ala Gln Lys Pro Glu Pro Asn Ile Phe Lys Pro Ser Ser
                          245 250 255
          Ile Ile Phe Phe Asp Glu Phe Lys Asn Thr Leu Ser Glu Asp Val Glu
                      260 265 270
          Arg Leu His Lys Asp Pro Lys Leu Tyr Leu Asp Phe Ile Ser Gln Asn
                  275 280 285
          Pro Phe Gln Asp Thr Ala Ala Glu Tyr Ile Ile Gln Thr Ile Ser Asn
              290 295 300
          Leu Glu Leu Lys Leu Lys Glu Ile Ile Asn Gln Ala
          305 310 315
           <![CDATA[ <210> 125]]>
           <![CDATA[ <211> 343]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Vibrio butyricum sp. TB]]>
           <![CDATA[ <400> 125]]>
          Met Asn Ile Ile His Phe Tyr Ala Arg Tyr Leu Arg Glu Ser His Asn
          1 5 10 15
          Trp Asn Arg Glu Arg Glu Val Thr Arg Asn Gly Val Met Thr Phe Ala
                      20 25 30
          Asn Trp Trp Arg Glu Asp Pro His Lys Asn Trp Phe Ala Arg Phe Ile
                  35 40 45
          Asp Ala Gly Ser Lys Asp Pro Glu Arg Arg Ile Arg Phe Tyr Ser Ile
              50 55 60
          Phe Gly Pro Tyr Ser Lys Leu Lys Glu Asp Phe Asp Gly Ala Lys Ile
          65 70 75 80
          Phe Phe Ser Gly Glu Asn Leu Glu Gln Pro Val Tyr His Arg Ile Leu
                          85 90 95
          Lys Thr Asp Pro Ile Glu Asp Arg Ile Trp Ala Asp Arg Arg Lys Leu
                      100 105 110
          Tyr Gly Asn Tyr Gly Ala Gly Asp Val Asp Leu Ala Ile Gly Phe Gly
                  115 120 125
          Asn Arg Glu Glu Asp Ser Leu Met Gly Phe Glu Gly Ser Arg Lys Thr
              130 135 140
          Lys Tyr Ile Arg Phe Pro Leu Trp Leu Thr Tyr Val Phe Asp Pro Asp
          145 150 155 160
          Cys Thr His Asp Asp Ile Lys Arg Thr Ile Asp Glu Ile Asn Ala Val
                          165 170 175
          Arg Ser Thr Gly Arg Lys Asp Thr Leu Leu Leu Ala Ser His Asp Phe
                      180 185 190
          Trp Gly Thr Arg Ser Asp Ile Leu Lys Ser Leu Glu Gly Val Cys Asp
                  195 200 205
          Val Ser Ile Ala Gly Lys Trp Arg Asn Asn Thr Lys Glu Leu Trp Glu
              210 215 220
          Asp Tyr Asn Asn Asp Lys Asn Lys Tyr Leu Ser Glu Phe Lys Phe Asn
          225 230 235 240
          Ile Cys Pro Glu Asn Val Asp Ala Pro Gly Tyr Val Thr Glu Lys Ile
                          245 250 255
          Phe Asp Ala Phe Lys Cys Gly Ala Ile Pro Ile Tyr Gln Gly Cys Leu
                      260 265 270
          Gly Lys Pro Glu Pro Asp Val Ile Asn Thr Asp Ala Val Leu Leu Trp
                  275 280 285
          Asp Phe Asp Gly Asp Asn Ser Asp Thr Ile Ser Leu Ile Lys Lys Leu
              290 295 300
          Asn Ser Asp Asn Val Tyr Tyr Asp Asn Phe Val Ser Gln Pro Lys Phe
          305 310 315 320
          Lys Pro Asp Ala Ala Glu Tyr Val Val Ala Cys Met Asp Glu Leu Arg
                          325 330 335
          Arg Ser Phe Asp Gln Leu Ile
                      340
           <![CDATA[ <210> 126]]>
           <![CDATA[ <211> 352]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Pyrromonas catarrhalis]]>
           <![CDATA[ <400> 126]]>
          Met Leu Ala Pro Tyr Lys Ser Pro Ile Phe Val Pro Ile Tyr Asp Thr
          1 5 10 15
          Lys Ala Met Asn Pro Pro Thr Lys Gln Pro Leu Arg Glu Arg Leu His
                      20 25 30
          Met Met Arg Arg Arg Asn Arg Ile Arg Lys Arg Ser Val Ile Ala Leu
                  35 40 45
          Ile Lys Ser His Leu Asp Ser Ser Arg Tyr Gln Asp Tyr Asn Trp Trp
              50 55 60
          Asp Ser His Ala Ser Thr Phe Trp Leu Pro Arg Phe Ile Asp Leu His
          65 70 75 80
          Leu Glu Pro Lys Lys Arg Ile Asn Leu Phe Ser Cys Phe Gln Asn Pro
                          85 90 95
          Leu Met Leu Ile Arg Tyr Tyr Lys Gly Val Lys Ile Phe Leu Ser Gly
                      100 105 110
          Glu Asn Leu Ala Asn Asn Glu His Phe Gly Phe His Pro Arg Met Leu
                  115 120 125
          Asp His Arg Ile Asn Glu Val Asp Leu Ala Leu Gly Phe Glu Phe Arg
              130 135 140
          Lys Asp Pro Lys Tyr Tyr Arg Phe Pro Leu Trp Ile Tyr Gln Asn Glu
          145 150 155 160
          Phe Ile Ser Pro Ser Ala Ser Leu Glu Asp Ile Arg Ala Leu Leu Glu
                          165 170 175
          Gln Ile Asn Asp Pro Ser Thr Arg Arg Ser Thr Gly Arg Ser Arg Phe
                      180 185 190
          Ile Gly Gln Ile Ser Ser His Asp Lys Gly Gly Met Arg Gly Arg Leu
                  195 200 205
          Ile Asp Leu Leu Asn Pro Ile Gly Gln Ile Asp Cys Ala Gly Lys Phe
              210 215 220
          Arg His Asn Thr Asp Glu Leu Leu Glu Val Tyr Gly Asp Asp Lys Phe
          225 230 235 240
          Lys Tyr Leu Ala Asn Tyr Arg Phe Asn Leu Cys Pro Glu Asn Ser Leu
                          245 250 255
          Gly Glu Gly Tyr Ile Thr Glu Lys Val Phe Asp Ser Ile Arg Ala Gly
                      260 265 270
          Cys Ile Pro Ile Tyr Trp Gly Ala Tyr Leu Glu Pro Gly Ile Leu Asn
                  275 280 285
          Pro Lys Ala Ile Leu Arg Phe Glu Glu Gly Lys Glu Gln Glu Phe Tyr
              290 295 300
          Asn Arg Val Lys Glu Leu Trp Glu Asn Glu Ala Ala Tyr Glu Gln Phe
          305 310 315 320
          Ile Leu Glu Pro Pro Phe Val Glu Gly Ala Ala Glu Arg Ile Trp Glu
                          325 330 335
          Ile Leu Gln Gly Leu Arg Glu Arg Leu Ala Pro Leu Val Glu Glu Gly
                      340 345 350
           <![CDATA[ <210> 127]]>
           <![CDATA[ <211> 343]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Vibrio cellolyticus butyricum]]>
           <![CDATA[ <400> 127]]>
          Met Asn Ile Ile His Phe Tyr Ala Arg Tyr Leu Arg Glu Ser His Asn
          1 5 10 15
          Trp Asn Arg Glu Arg Glu Val Thr Arg Asn Gly Val Met Thr Phe Ala
                      20 25 30
          Asn Trp Trp Arg Glu Asp Pro His Lys Asn Trp Phe Ala Arg Phe Ile
                  35 40 45
          Asp Ala Gly Asn Lys Asp Pro Glu Arg Arg Ile Arg Phe Tyr Ser Ile
              50 55 60
          Phe Gly Pro Tyr Ser Lys Leu Lys Glu Asp Phe Asp Gly Ala Lys Ile
          65 70 75 80
          Phe Phe Ser Gly Glu Asn Leu Glu Gln Pro Val Leu His Arg Ile Leu
                          85 90 95
          Lys Thr Asp Pro Ile Glu Asp Arg Ile Trp Ala Asp Arg Arg Lys Leu
                      100 105 110
          Tyr Gly Asn Tyr Gly Ala Gly Glu Val Asp Leu Ala Ile Gly Phe Gly
                  115 120 125
          Asn Arg Glu Glu Asp Ser Leu Leu Gly Phe Glu Gly Ser Arg Lys Thr
              130 135 140
          Lys Tyr Ile Arg Phe Pro Leu Trp Leu Thr Tyr Val Phe Asp Pro Asp
          145 150 155 160
          Cys Thr His Asp Asp Ile Lys Arg Thr Ile Asp Glu Ile Asn Ala Val
                          165 170 175
          Arg Ser Thr Gly Arg Lys Asp Thr Leu Leu Leu Ala Ser His Asp Phe
                      180 185 190
          Trp Gly Thr Arg Ser Asp Ile Leu Lys Ser Leu Glu Gly Val Cys Asp
                  195 200 205
          Ile Ser Ile Ala Gly Lys Trp Arg Asn Asn Thr Lys Glu Leu Trp Glu
              210 215 220
          Asp Tyr Asp Asn Asp Lys Asn Lys Tyr Leu Ser Glu Phe Lys Phe Asn
          225 230 235 240
          Ile Cys Pro Glu Asn Val Asp Ala Pro Gly Tyr Val Thr Glu Lys Ile
                          245 250 255
          Phe Asp Ala Phe Lys Cys Gly Ala Ile Pro Ile Tyr Gln Gly Cys Leu
                      260 265 270
          Gly Lys Pro Glu Pro Asn Val Ile Asn Thr Asp Ala Val Leu Leu Trp
                  275 280 285
          Asp Phe Asp Gly Asp Asn Ser Asp Thr Ile Ala Leu Ile Lys Lys Leu
              290 295 300
          Asn Ser Asp Asn Val Tyr Tyr Asp Asn Phe Val Ser Gln Pro Lys Phe
          305 310 315 320
          Lys Pro Asp Ala Ala Glu Tyr Val Val Ala Cys Met Asp Glu Leu Arg
                          325 330 335
          Arg Ser Phe Asp Arg Leu Ile
                      340
           <![CDATA[ <210> 128]]>
           <![CDATA[ <211> 366]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Rice straw rhizobia]]>
           <![CDATA[ <400> 128]]>
          Met Leu Cys Pro Asp Ala Trp Gly Trp Leu Ser Ile Ser Ala Lys Ala
          1 5 10 15
          Leu Val Asn Pro Arg Glu Ala Thr Lys Ser Glu Ala Gly Gly Arg Tyr
                      20 25 30
          Glu Gln Gly Thr Ala Met Lys Thr Val Arg Val Trp Tyr Thr Asp Phe
                  35 40 45
          Trp Pro Asp Phe Lys Pro Glu Asp Leu Phe Leu Leu Glu Gly Arg Ser
              50 55 60
          Gly Asp Val Glu Val Val Leu Asp Pro Lys Ser Pro Asp Val Leu Phe
          65 70 75 80
          Tyr Ser Cys Phe Asp Thr Val His Arg Thr Tyr Asp Cys Val Arg Val
                          85 90 95
          Phe Tyr Thr Gly Glu Asn Val Arg Pro Asp Phe Asn Val Cys Asp Tyr
                      100 105 110
          Gly Ile Gly Phe Asp His Met Ser Phe Gly Asp Arg Tyr Leu Arg His
                  115 120 125
          Pro Phe Tyr Leu Glu Asp Leu Ala Leu Ala Glu Gln Val Arg Asp Arg
              130 135 140
          Ala Pro Val Asp Ala Arg Leu Val Glu Gly Arg Gly Phe Cys Thr Phe
          145 150 155 160
          Ile Tyr Ser Asn Gly Arg Ala Asp Pro Val Arg Asp Gly Phe Phe His
                          165 170 175
          His Leu Ser Arg His Arg Pro Val Asn Ser Met Gly Arg His Leu Arg
                      180 185 190
          Asn Asp Asp Ser Phe Ala Arg Met Thr Gly Leu Lys Pro Val Pro Ala
                  195 200 205
          Lys Leu Lys Val Met Gly Asp Phe Asn Phe Ala Ile Ala Phe Glu Asn
              210 215 220
          Ser Gly Thr Pro Gly Tyr Thr Thr Glu Lys Ile Phe His Ala Phe Ala
          225 230 235 240
          Ala Arg Cys Ile Pro Ile Tyr Trp Gly Asn Pro Leu Val Ala Leu Asp
                          245 250 255
          Phe Asn Pro Arg Ala Phe Val Asn Arg His Asp Phe Pro Asp Asp Asp
                      260 265 270
          Ser Cys Ile Ala His Val Met Glu Leu Ala Arg Asp Pro Gln Arg Met
                  275 280 285
          Val Ala Met Leu Asn Glu Pro Val Phe Ala Glu Gly Asn Asp Pro Ser
              290 295 300
          Leu Arg Arg Gln Gln Leu Ala Asp Phe Ile Asn His Ile Phe His Gln
          305 310 315 320
          Ala Pro Glu Glu Ala Arg Arg Arg Thr Arg Tyr Gly Tyr Ala Pro Val
                          325 330 335
          Tyr Val Arg Arg Asn Glu Pro Arg Gln Arg Ser Trp Phe Ser Lys Leu
                      340 345 350
          Asn Met Lys Arg Lys Ala Trp Arg Lys Ala Met Arg Ser Arg
                  355 360 365
           <![CDATA[ <210> 129]]>
           <![CDATA[ <211> 337]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Levania xylanase]]>
           <![CDATA[ <400> 129]]>
          Met Phe Arg Pro Leu Lys Gln Pro Val Lys Asn Val Val Ser Gly Leu
          1 5 10 15
          Ile Gly Glu Glu Arg Leu Thr Ala Tyr Asn Arg Tyr Ala Arg Gln Thr
                      20 25 30
          Ala Gly Ala Tyr Tyr Gln His Phe Leu Leu Arg Cys Tyr Gly Arg Gly
                  35 40 45
          Phe Tyr Asn Phe Trp Arg Val Pro Ser Phe Arg Asn Thr Trp Phe Trp
              50 55 60
          Arg Phe Leu Asp Arg Pro Arg Leu Ala Lys Leu Ser Lys Asp Arg Arg
          65 70 75 80
          Ile Ala Phe Ile Ser Val Phe Gly Asp Pro Arg Leu Ile Ser Arg Ile
                          85 90 95
          Asn Ala Asp Leu Arg Val Phe Phe Thr Gly Glu Asn Leu Asp Phe His
                      100 105 110
          Pro Ser Tyr Arg Asp His Leu Phe Gly Ser Val Asp Leu Ser Leu Gly
                  115 120 125
          Phe Asp Gln Leu Gln Arg Ser Asp Tyr Leu Arg Phe Pro Ile Trp Leu
              130 135 140
          Leu Leu Cys Phe Asp Pro Gly Met Gln Leu Ala Asp Ile Thr Glu Arg
          145 150 155 160
          Leu Arg Gln Trp Asp Arg Asn Arg Leu Asp Leu Leu Gln Arg Pro Ala
                          165 170 175
          Met Val Ala Ser Leu Val Ala Ser His Asp Pro Ser Gly Ser Arg Arg
                      180 185 190
          Arg Ile Ala Asp Leu Phe Asp Gln Val Gly Thr Val Arg Tyr Gly Gly
                  195 200 205
          Ala Phe Arg Asn Val Gly Glu Arg Val Pro Pro Gly Gln Ala Tyr Lys
              210 215 220
          His Arg Phe Ile Arg Gln Tyr Pro Phe His Ile Cys Pro Glu Asn Ser
          225 230 235 240
          Gln Arg Pro Gly Tyr Thr Thr Glu Lys Leu Phe Glu Ala Val Ala Ala
                          245 250 255
          Gly Cys Ile Pro Ile Tyr Arg Gly Ala Glu Gly Ala Val Glu Pro Asp
                      260 265 270
          Ile Phe Asn Gln Asp Ala Ile Ile Ala Tyr Ser Pro Gly Glu Glu Thr
                  275 280 285
          Val Phe Arg Gln Arg Leu Arg Ser Leu Arg Asp Asp Pro His Gln Leu
              290 295 300
          Arg Asp Leu Arg Gly Ile Pro Pro Phe Arg Pro Asp Ala Ala Glu His
          305 310 315 320
          Ile Tyr Asn Phe Tyr Leu Arg Leu Glu Glu Lys Leu Phe Asp Leu Leu
                          325 330 335
          Gly
           <![CDATA[ <210> 130]]>
           <![CDATA[ <211> 327]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Homo sapiens]]>
           <![CDATA[ <400> 130]]>
          Met Ile Arg Asn Ile Lys Asn Lys Leu Lys Asn Thr Ile His Leu Leu
          1 5 10 15
          Lys Val Tyr Ile Asn Ser Gly Gln Ile Asn Phe Tyr Asn Phe Trp Thr
                      20 25 30
          Glu Asn Asn Ile Asp Asp Tyr Trp Leu Tyr Arg Phe Val Lys Ser Arg
                  35 40 45
          Lys Ile Asn Asn Lys Ile Ser Phe Phe Ser Val Phe Gly Lys Arg Phe
              50 55 60
          Ile Val Asp Tyr Ser Phe Gly Val Gly Lys Lys Val Phe Phe Thr Gly
          65 70 75 80
          Glu Asn Leu Thr Glu Thr Asn Ile Phe Phe Leu Gly Tyr Ala Pro Lys
                          85 90 95
          Tyr Lys Asn His Cys Leu Asn Ser Val Asp Leu Ser Leu Gly Phe Asp
                      100 105 110
          Tyr Ile Asp His Pro Lys Tyr Met Arg Phe Pro Leu Trp Ile Lys Tyr
                  115 120 125
          Leu Thr Asn Gly Thr Glu Gln Ser Ile Asn Asp Ile Glu Glu Lys Ile
              130 135 140
          Asn Asn Ile Asn Asn Pro Arg His Arg Leu Ser Lys Gly Arg Asn Tyr
          145 150 155 160
          Phe Ala Ser Leu Ile Ser Arg His Asp Glu Gly Gly Thr Arg Met Lys
                          165 170 175
          Leu Ile Asn Leu Leu Asn Pro Ile Asn Lys Val Val Cys Ala Gly Thr
                      180 185 190
          Phe Met Asn Asn Thr Asn Asn Leu Lys Glu Lys Phe Asn Asp Asn Lys
                  195 200 205
          Leu Glu Phe Leu Lys Gln Phe Lys Phe Asn Ile Cys Pro Glu Asn Ser
              210 215 220
          Arg Ala Asn Gly Tyr Ile Thr Glu Lys Ile Phe Glu Ala Ile Ile Ser
          225 230 235 240
          Gly Cys Ile Pro Ile Tyr Tyr Gly Gly Arg Asp Thr Ser Asn Thr Asn
                          245 250 255
          Glu Ala Lys Gln Ser Ile Glu Pro Glu Ile Leu Asn Pro Glu Ala Phe
                      260 265 270
          Ile Phe Tyr Asn Gly Asn Asn Glu Glu Glu Val Phe Ala Lys Val Lys
                  275 280 285
          Glu Leu Trp Glu Asn Gln Ser Ala Tyr Glu Glu Phe Cys Lys Ile Pro
              290 295 300
          Pro Phe Lys Glu Asn Ala Ala Lys Val Ile Trp Ser Lys Leu Gln Glu
          305 310 315 320
          Leu Glu Asp Arg Leu Lys Lys
                          325
           <![CDATA[ <210> 131]]>
           <![CDATA[ <211> 429]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Bisgard taxon 44 strain B96_3]]>
           <![CDATA[ <400> 131]]>
          Met Ala Asp Ile Asn His Cys Gly Tyr Trp Leu Gly Phe Ser Leu Glu
          1 5 10 15
          Lys Gln Lys Ser Ile Phe Leu Tyr His Lys Phe Asp Leu Gln Lys Leu
                      20 25 30
          Pro His Asn Leu Phe Leu Val Ser Gly Phe Leu Arg Arg Phe Trp Thr
                  35 40 45
          Gln Ser Glu Leu Glu Arg Arg Ala Ser Gly Tyr Gln Glu Lys Leu Gly
              50 55 60
          Trp Ala Pro Leu Leu Tyr His Gly Tyr Pro Tyr Asn Ala Ala Ser Cys
          65 70 75 80
          Leu Gly Ile Pro Tyr Lys Leu Lys His Gly Thr Ser Glu Arg Ser Phe
                          85 90 95
          Tyr Tyr Asp Gln Glu His Ser Leu Glu Phe Ala Phe Ser Glu Asn Ser
                      100 105 110
          Gln Asp Ile Tyr Ala Asp Tyr Leu Asp Leu Ala Leu Gly Ile Tyr Leu
                  115 120 125
          Pro Glu Val Ile Ser Thr Arg Tyr Cys Asn Tyr Tyr His Ala Pro Asn
              130 135 140
          Tyr Leu Val Ser Gln Ser Val Gly Leu Asn Thr Leu Tyr Gly Leu Pro
          145 150 155 160
          Thr Tyr Ala Gln Val Glu Ala Trp Ile Glu Ser Phe Glu Gln Ala Arg
                          165 170 175
          Arg Gln Ala Glu Val Phe Lys Arg Pro Ala Leu Ser Val Cys Ile Ser
                      180 185 190
          Arg His Asp Ser Leu Ser Gly Leu Arg Gly Val Arg Gly Leu Ile Gly
                  195 200 205
          Asp Leu Phe Thr Glu Val Thr Lys Asp Leu His Glu Ala Pro Val Phe
              210 215 220
          Tyr Ala Gly Tyr Phe Arg Asn Asn Thr Ser Asp Leu His Glu His Phe
          225 230 235 240
          Asn Asp Asp Lys Ile Ala Tyr Gly Lys Asn Phe Val Phe Met Ile Cys
                          245 250 255
          Pro Glu Asn Ser Tyr Cys Pro Gly Tyr Val Ser Glu Lys Ile Leu Glu
                      260 265 270
          Ala Phe Ala Ser Gly Cys Ile Pro Ile Tyr Trp Gly Gly Ile Glu Ser
                  275 280 285
          Glu Leu Asp Tyr Phe Asn Glu Lys Ala Phe Val Tyr Phe Asp Pro Arg
              290 295 300
          Lys Pro Lys Glu Phe Gln Glu Arg Ile Arg Glu Leu Val Tyr Asp Gln
          305 310 315 320
          Gly Lys Leu Glu Glu Met Ile Ala Gln Pro Cys Phe Leu Pro Gly Ala
                          325 330 335
          Ala Ala Arg Ile Tyr Leu Arg Tyr Ile Tyr Pro Ile Ala Val Ala Phe
                      340 345 350
          Ser Asn Leu Met Asp Gly Leu Asn Pro Tyr Gln Leu Leu Lys Glu Pro
                  355 360 365
          Val Asp Thr Thr Leu Glu Gln Glu Lys Ala Lys Ala Glu Ala Val Leu
              370 375 380
          Ala Gln Leu Gly Leu Val Glu Asn Lys Tyr Gln Gly Leu Asp Arg Leu
          385 390 395 400
          Pro Phe Lys Tyr Tyr Val Asp Phe Glu Gln Tyr Asn Glu Glu Glu Lys
                          405 410 415
          Gln Arg Leu Glu Tyr Phe Val Arg His Tyr Asn Ser Lys
                      420 425
           <![CDATA[ <210> 132]]>
           <![CDATA[ <211> 327]]>
           <![CDATA[ <212> PRT]]>
           <![CDATA[ <213> Corynebacterium xylomethyl]]>
           <![CDATA[ <400> 132]]>
          Met Ile Phe Lys Ala Leu Glu Phe Phe Thr Arg Arg Ser Pro Gly Ala
          1 5 10 15
          Arg Thr Pro Ala Thr Asp Arg Gln Ile Ala Ser Glu Ala Leu Ala Ile
                      20 25 30
          Cys Ala Val Asp Phe Trp Pro Ala Phe Ser Leu Ser Gly Gly Phe Trp
                  35 40 45
          Asn Phe Val Leu Thr Glu Ser Phe Gly Gln Phe Arg Val Val Asp Glu
              50 55 60
          Ala Ala Asp Ala Asp Ile Val Leu Ala Ser Val Phe Pro His Glu Lys
          65 70 75 80
          Ala Gln Phe Pro Glu Lys Thr Ile Ala Ile Ile Trp Glu Asn Ile Arg
                          85 90 95
          Pro Asn Tyr Glu Phe Tyr Arg Phe Ser Ile Ser Ser Asp Phe Asp Ala
                      100 105 110
          Tyr Gln Gly Arg Asn Cys Arg Val Pro Asn Trp Tyr Glu Glu Leu Val
                  115 120 125
          Trp Asn Glu Ser Phe Arg Gly Pro Lys Ala Ala Phe Ala Gly Ala Met
              130 135 140
          His Gly His Gly His Glu Glu Arg Val Glu Ile Glu Arg Leu Leu Ala
          145 150 155 160
          Pro Gly Glu Ala Glu Pro Val Val Arg Ser Lys Phe Cys Cys Leu Val
                          165 170 175
          Thr Ser Tyr Arg Glu Pro Tyr Arg Ala Leu Ala Val Glu Ala Leu Ser
                      180 185 190
          Gln Ile Gly Pro Val Asp Ile Phe Gly Asn Val Ala Gly Ala Pro Leu
                  195 200 205
          Leu Gln Ser Lys Tyr Glu Ala Leu Arg Asp Tyr Arg Phe Asn Leu Cys
              210 215 220
          Phe Glu Asn Ser Ile Phe Pro Gly Tyr Tyr Thr Glu Lys Ala Leu Gln
          225 230 235 240
          Ala Trp Ala Ala Gly Cys Ile Pro Leu Tyr Phe Ser Asp Pro Tyr Phe
                          245 250 255
          Ser Ala Asp Phe Asn Pro Lys Ala Ile Ile Asn Arg Ile Asp Phe Arg
                      260 265 270
          Glu Leu Ala Glu Phe Val Glu Ala Val Arg Arg Val Asn Asp Ser Pro
                  275 280 285
          Asp Leu Met Ala Gln Phe His Arg Glu Pro Leu Leu Thr Ser Arg Pro
              290 295 300
          Asn Leu Asp Ser Val Thr Ala Phe Leu Arg Gly Ala Ala Lys Ala Ile
          305 310 315 320
          Thr Gly Arg Asp Ile Gly Gly
                          325
          
      

Claims (61)

A method for producing 2',3-difucosyllactose (DiFL) comprising the steps of: i) Provide at least two fucosyltransferases, preferably at least one α-1,2-fucosyltransferase and at least one α-1,3-fucosyltransferase, and GDP-fucosyltransferase sugar donors, wherein the fucosyltransferases are capable of transferring fucose residues from the GDP-fucose donor to one or more acceptors, and, ii) subjecting the fucosyltransferase and GDP-fucose donor to a mixture comprising acceptor lactose and possibly one or more other acceptors on the fucosyltransferase catalyzing fucose residues Contacting under conditions that transfer from the GDP-fucose donor to the acceptor(s), thereby producing DiFL and possibly 2'FL and/or 3FL, iii) Preferably, at least one of the 2'FL, 3-FL and DiFL is separated, more preferably, all of the 2'FL, 3-FL and DiFL are separated. The method of claim 1, wherein the DiFL is present in the reaction mixture at a purity of at least 80% as measured by the total amount of 2'FL, 3-FL and DiFL produced in the mixture. The method of any one of claims 1 or 2, wherein the fucosyltransferases and the GDP-fucose are provided in a cell-free system. The method of any one of claims 1 or 2, wherein the fucosyltransferases and the GDP-fucose are metabolically engineered by cells, preferably single cells, more preferably for the production of DiFL or a mixture of 2'FL, 3-FL and DiFL provided by individual cells. The method of any one of claims 1 to 4, wherein the other acceptor(s) is selected from a list comprising 2'FL and 3-FL. The method of any one of claims 1, 2, 4 or 5, the method comprising the following steps: a. providing a cell, preferably a single cell, wherein the cell expresses at least two fucosyltransferases, preferably at least one alpha-1,2-fucosyltransferase and at least one alpha-1,3-peptidase Fluorosyltransferases capable of transferring fucose residues from a GDP-fucose donor to one or more of the acceptor lactose and possibly the other acceptor(s) , and the cell is capable of synthesizing GDP-fucose, wherein the GDP-fucose is a donor for the fucosyltransferases, and b. culturing the cells under conditions that allow expression of the fucosyltransferases and synthesis of the GDP-fucose, c. Preferably, at least one of the 2'FL, 3-FL or DiFL is isolated from the culture, more preferably all of the 2'FL, 3-FL and DiFL are isolated from the culture. The method of any one of claims 4 to 6, wherein the cell exhibits: a. At least one alpha-1,2-fucosyltransferase that (i) comprises the polypeptide sequence of any one of SEQ ID NOs: 16 to 110, or (ii) is SEQ ID NO: 16-110 A functional homologue, variant or derivative of any one of ID NOs 16 to 110 which is the same as the alpha-1,2-rock having SEQ ID NOs 16 to 110 The full length of any of the glucosyltransferase polypeptides has at least 80% overall sequence identity and has alpha-1 for lactose and possibly any of the other receptors 2'FL and/or 3FL ,2-fucosyltransferase activity, or (iii) is a functional fragment of any one of SEQ ID NOs 16 to 110 and has alpha-1,2-fucosyltransferase activity, or (iv) Comprising a polypeptide comprising or consisting of an amino acid sequence having at least 80% sequence identity to the full-length amino acid sequence of any one of SEQ ID NOs 16 to 110 and having alpha-1,2-fucoid glycosyltransferase activity, and b. at least one α-1,3-fucosyltransferase, the fucosyltransferase (i) comprising as any one of SEQ ID NOs: 111 to 132, preferably SEQ ID NOs: 113 to any one of 132, more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NO: 113 , any one of 114, 118, 119, 120, 121 or 122, even better the polypeptide sequence of SEQ ID NO: 113 or 114, the best SEQ ID NO: 114, or (ii) the polypeptide sequence of SEQ ID NO: 111 to 114 any one of 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or Any one of 132, even better any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: The functional homologue, variant or derivative of 114, which is related to any one of SEQ ID NOs 111 to 132, preferably SEQ ID NOs: 113 to 132, more preferably Any of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NOs: 113, 114, 118, 119, 120, The full length of any one of 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: 114 of any one of the alpha-1,3-fucosyltransferase polypeptides has At least 80% overall sequence identity and alpha-1,3-fucosyltransferase activity on lactose and possibly on any of the (or) other receptors 2'FL and/or 3FL, or (iii) ) is any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128 , any one of 129, 130, 131 or 132, even more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even more preferably SEQ ID NO: 113 or 114 , the best functional fragment of SEQ ID NO: 114 with alpha-1,3-fucosyl transfer Enzymatic activity, or (iv) comprising a polypeptide comprising and any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114 , any of 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably any of SEQ ID NOs: 113, 114, 118, 119, 120, 121 or 122 or, even more preferably SEQ ID NO: 113 or 114, the full-length amino acid sequence of SEQ ID NO: 114 has or consists of an amino acid sequence having at least 80% sequence identity and has α-1,3- Fucosyltransferase activity. The method of claim 7, wherein the cell exhibits: a. at least one α-1,2-fucosyltransferase comprising as SEQ ID NOs: 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to the polypeptide sequence of any one of 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105, or (ii) is SEQ ID NO 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 A functional homologue, variant or derivative of any of the functional homologues, variants or derivatives having SEQ ID NOs 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52 , 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 of any of the α-1,2-fucosyltransferase polypeptides has at least 80% overall sequence identity over its full length and has alpha-1,2-fucosyltransferase activity on lactose and possibly on any of the other receptors 2'FL and/or 3FL(s), or (iii) is SEQ ID NO 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to A functional fragment of any one of 100, 102, 103 or 105 and having α-1,2-fucosyltransferase activity, or (iv) comprising a polypeptide comprising and SEQ ID NOs 19 to 21, 23 to any of 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 A full-length amino acid sequence having or consisting of an amino acid sequence having at least 80% sequence identity and having alpha-1,2-fucosyltransferase activity, and b. at least one α-1,3-fucosyltransferase, the fucosyltransferase (i) comprising as any one of SEQ ID NOs: 111 to 132, preferably SEQ ID NOs: 113 to any one of 132, more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NO: 113 , any one of 114, 118, 119, 120, 121 or 122, even better the polypeptide sequence of SEQ ID NO: 113 or 114, the best SEQ ID NO: 114, or (ii) the polypeptide sequence of SEQ ID NO: 111 to 114 any one of 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or Any one of 132, even better any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: The functional homologue, variant or derivative of 114, which is related to any one of SEQ ID NOs 111 to 132, preferably SEQ ID NOs: 113 to 132, more preferably Any of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NOs: 113, 114, 118, 119, 120, The full length of any one of 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: 114 of any one of the alpha-1,3-fucosyltransferase polypeptides has At least 80% overall sequence identity and alpha-1,3-fucosyltransferase activity on lactose and possibly on any of the (or) other receptors 2'FL and/or 3FL, or (iii) ) is any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128 , any one of 129, 130, 131 or 132, even more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even more preferably SEQ ID NO: 113 or 114 , the best functional fragment of SEQ ID NO: 114 with alpha-1,3-fucosyl transfer Enzymatic activity, or (iv) comprising a polypeptide comprising and any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114 , any of 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably any of SEQ ID NOs: 113, 114, 118, 119, 120, 121 or 122 or, even more preferably SEQ ID NO: 113 or 114, the full-length amino acid sequence of SEQ ID NO: 114 has or consists of an amino acid sequence having at least 80% sequence identity and has α-1,3- Fucosyltransferase activity. The method of claim 7, wherein the cell exhibits: a. At least one α-1,2-fucosyltransferase, the fucosyltransferase (i) comprising as SEQ ID NO: 16 to 19 any one, preferably SEQ ID NO 16 or 19 , more preferably the polypeptide sequence of SEQ ID NO 16, or (ii) any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 or 19, more preferably a functional homologue, variation of SEQ ID NO 16 body or derivative, the functional homologue, variant or derivative with the alpha-1,2-fucoid having SEQ ID NO 16 to 19, preferably SEQ ID NO 16 or 19, more preferably SEQ ID NO 16 The full length of any of the glycosyltransferase polypeptides has at least 80% overall sequence identity and has alpha-1 for lactose and possibly any of the other receptors 2'FL and/or 3FL(s), 2-fucosyltransferase activity, or (iii) is any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 or 19, more preferably a functional fragment of SEQ ID NO 16 and having alpha-1 , 2-fucosyltransferase activity, or (iv) comprising a polypeptide comprising any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 or 19, more preferably SEQ ID NO 16 A full-length amino acid sequence having or consisting of an amino acid sequence having at least 80% sequence identity and having alpha-1,2-fucosyltransferase activity, and b. at least one α-1,3-fucosyltransferase, the fucosyltransferase (i) comprising as any one of SEQ ID NOs: 111 to 132, preferably SEQ ID NOs: 113 to any one of 132, more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NO: 113 , any one of 114, 118, 119, 120, 121 or 122, even better the polypeptide sequence of SEQ ID NO: 113 or 114, the best SEQ ID NO: 114, or (ii) the polypeptide sequence of SEQ ID NO: 111 to 114 any one of 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or Any one of 132, even better any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: The functional homologue, variant or derivative of 114, which is related to any one of SEQ ID NOs 111 to 132, preferably SEQ ID NOs: 113 to 132, more preferably Any of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NOs: 113, 114, 118, 119, 120, The full length of any one of 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: 114 of any one of the alpha-1,3-fucosyltransferase polypeptides has At least 80% overall sequence identity and alpha-1,3-fucosyltransferase activity on lactose and possibly on any of the (or) other receptors 2'FL and/or 3FL, or (iii) ) is any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128 , any one of 129, 130, 131 or 132, even more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even more preferably SEQ ID NO: 113 or 114 , the best functional fragment of SEQ ID NO: 114 with alpha-1,3-fucosyl transfer Enzymatic activity, or (iv) comprising a polypeptide comprising and any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114 , any of 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably any of SEQ ID NOs: 113, 114, 118, 119, 120, 121 or 122 or, even more preferably SEQ ID NO: 113 or 114, the full-length amino acid sequence of SEQ ID NO: 114 has or consists of an amino acid sequence having at least 80% sequence identity and has α-1,3- Fucosyltransferase activity. The method of any one of claims 4 to 9, wherein the cell produces lactose for the synthesis of any of the 2'FL, 3-FL and DiFL, preferably the cell further produces the one or more other receptors at least one of the bodies. The method of any one of claims 1 to 10, wherein the method comprises using one or more precursors for the synthesis of any of the 2'FL, 3-FL and DiFL. The method of any one of claims 1 to 11, wherein the precursor(s) and/or acceptor are fully converted to any of the 2'FL, 3-FL and DiFL. The method of any one of claims 4 to 12, wherein the cell is modified in the expression or activity of at least one of the fucosyltransferases. The method of any one of claims 4 to 13, wherein the cell is a metabolically engineered cell. The method of claim 14, wherein the cell comprises multiple copies of the same coding DNA sequence encoding a protein. The method of any one of claims 4 to 15, wherein the cell is modified to produce GDP-fucose. The method of any one of claims 4 to 16, wherein the cell is modified for enhanced GDP-fucose production compared to an unmodified precursor cell, and wherein the modification is selected from the group consisting of Group: UDP-glucose: undecaprenyl-phosphoglucose-1-phosphotransferase-encoding gene knockout, GDP-L-fucose synthase-encoding gene overexpression, GDP-mannose 4, Overexpression of a gene encoding 6-dehydratase, overexpression of a gene encoding mannose-1-phosphate guanosyltransferase, overexpression of a gene encoding phosphomannose mutase, or overexpression of a gene encoding mannose-6-phosphate isomerase Overexpression of genes. The method of any one of claims 4 to 17, wherein the cells are resistant to lactose killing when grown in an environment where lactose is combined with one or more other carbon sources. The method of any one of claims 4 to 18, wherein the cell produces the DiFL or a mixture of the 2'FL, 3-FL and DiFL intracellularly and wherein the produced DiFL, 2'FL and/or 3-FL Some or substantially all of it remains intracellular and/or is excreted outside the cell via passive or active transport. The method of any one of claims 4 to 19, wherein the cell is further genetically modified for i) Modified expression of endogenous membrane proteins, and/or ii) the modified activity of the endogenous membrane protein, and/or iii) Representation of homologous membrane proteins, and/or iv) expression of heterologous membrane proteins, wherein the membrane protein participates in the secretion of any one of the 2'FL, 3-FL and DiFL from the mixture to the outside of the cell, preferably, the membrane protein participates in the secretion of the 2'FL from the mixture to the outside of the cell All of 2'FL, 3-FL and DiFL. The method of any one of claims 4 to 20, wherein the cell is further genetically modified for i) Modified expression of endogenous membrane proteins, and/or ii) the modified activity of the endogenous membrane protein, and/or iii) Representation of homologous membrane proteins, and/or iv) expression of heterologous membrane proteins, wherein the membrane protein is involved in the uptake of lactose and/or at least one other receptor 2'FL and/or 3FL for the synthesis of any of the 2'FL, 3-FL and DiFL. The method of any one of claims 20 or 21, wherein the membrane protein is selected from the list comprising: transporter, P-P-bond hydrolysis driven transporter, β-barrel pore proteins, transport proteins, putative transport proteins and phosphotransfer driven group translocation proteins, Preferably, these transport proteins comprise MFS transport proteins, carbohydrate efflux transport proteins and chelating ferritin export proteins, Preferably, the P-P-bond hydrolysis-driven transport proteins comprise ABC transport proteins and chelatin export proteins. The method of any one of claims 20 to 22, wherein the membrane protein improves the production of any of the 2'FL, 3-FL and DiFL and/or enables and/or enhances the 2'FL, 3-FL and DiFL - Outflow of either FL and DiFL. The method of any one of claims 4 to 23, wherein the cells are stably cultured in culture medium. The method of any one of claims 4 to 24, wherein the conditions include: (i) using a culture medium comprising at least one precursor and/or acceptor for the production of the DiFL or the mixture of 2'FL, 3-FL and DiFL, and/or (ii) adding to the medium at least one precursor and/or acceptor feed for the production of the DiFL or the mixture of 2'FL, 3-FL and DiFL. The method of any one of claims 4 to 25, comprising at least one of the following steps: i) using a medium comprising at least one precursor and/or acceptor; ii) adding to the medium in the reactor at least one precursor and/or acceptor feed, wherein the total reactor volume is in the range of 250 mL (milliliters) to 10.000 ^m3 (cubic meters), preferably in a continuous manner, and preferably such that the final the volume is not more than three times, preferably not more than twice, more preferably less than twice the volume of the medium before adding the precursor and/or acceptor feed; iii) adding at least one precursor to the medium in the reactor and /or acceptor feed, wherein the total reactor volume is in the range of 250 mL (milliliters) to 10.000 ^m3 (cubic meters), preferably in a continuous manner, and preferably such that the final volume of the medium does not exceed the addition Three times, preferably no more than twice, more preferably less than twice the volume of the medium before the precursor and/or acceptor feed, and wherein preferably the pH setting of the precursor and/or acceptor feed between 3 and 7, and wherein preferably, the temperature of the precursor and/or acceptor feed is maintained between 20°C and 80°C; iv) over 1 day, 2 days, 3 days, 4 days, Add at least one precursor and/or acceptor feed to the medium in a continuous manner with the aid of a feed solution over a 5 day time course; v) over a 1 day, 2 day, 3 day, 4 day, 5 day time course with the aid of at least one precursor and/or acceptor feed is added to the culture medium in a continuous manner in a feed solution, and wherein preferably the pH of the feed solution is set between 3 and 7, and wherein preferably the The temperature of the feed solution is maintained between 20°C and 80°C; the method produces a concentration in the final culture of at least 50 g/L, preferably at least 75 g/L, more preferably at least 90 g/L, more preferably at least 100 g/L g/L, more preferably at least 125 g/L, more preferably at least 150 g/L, more preferably at least 175 g/L, more preferably at least 200 g/L of DiFL or the mixture of 2'FL, 3-FL and DiFL any of the 2'FL, 3-FL and DiFL. The method of any one of claims 4 to 25, comprising at least one of the following steps: i) using a method comprising at least 50, more preferably at least 75, more preferably at least 100, more preferably at least 1 per liter of initial reactor volume 120. A culture medium containing at least 150 grams of lactose, more preferably, wherein the reactor volume is in the range of 250 mL to 10.000 m ³ (cubic meters); ii) adding to the culture medium a content of at least 50, more preferably, per liter of initial reactor volume Lactose feed of at least 75, more preferably at least 100, more preferably at least 120, more preferably at least 150 grams of lactose, wherein the reactor volume is in the range of 250 mL to 10.000 ^m3 (cubic meters), preferably in a continuous manner Add, and preferably such that the final volume of the medium does not exceed three times, preferably not more than twice, more preferably less than 2 times the volume of the medium before adding the lactose feed; iii) adding to the medium comprises an initial reaction per liter Lactose feed of at least 50, more preferably at least 75, more preferably at least 100, more preferably at least 120, more preferably at least 150 grams of lactose, wherein the reactor volume is in the range of 250 mL to 10.000 ^m3 (cubic meters) preferably in a continuous manner, and preferably such that the final volume of the medium is no more than three times, preferably no more than two times, more preferably less than two times the volume of the medium before the addition of the lactose feed, and preferably Preferably, the pH of the lactose feed is set between 3 and 7, and wherein preferably, the temperature of the lactose feed is maintained between 20°C and 80°C; iv) over 1 day, 2 days, 3 days, Lactose feed was added to the medium in a continuous manner with the aid of the feed solution over the course of 4 days, 5 days; v) over the course of 1 day, 2 days, 3 days, 4 days, 5 days with the aid of the feed solution Lactose feed is added to the culture medium in a continuous manner, and wherein the lactose feed solution has a concentration of 50 g/L, preferably 75 g/L, more preferably 100 g/L, more preferably 125 g/L, more preferably 150 g /L, better 175 g/L, better 200 g/L, better 225 g/L, better 250 g/L, better 275 g/L, better 300 g/L, better 325 g/L L, better 350 g/L, better 375 g/L, better 400 g/L, better 450 g/L, better 500 g/L, even better 550 g/L, best 600 g/L L; and wherein preferably the pH of the feed solution is set between 3 and 7, and wherein preferably the temperature of the feed solution is maintained between 20°C and 80°C; The resulting concentration in the final volume is at least 50 g/L, preferably at least 75 g/L, more preferably at least 90 g/L, more preferably at least 100 g/L, more preferably at least 125 g/L, more preferably at least 150 g/L L, preferably at least 175 g/L, More preferably at least 200 g/L of DiFL or any of the 2'FL, 3-FL and DiFL in the mixture of 2'FL, 3-FL and DiFL. The method of claim 27, wherein the lactose is fed by starting the culture at a concentration of at least 5 mM, preferably at a concentration of 30, 40, 50, 60, 70, 80, 90, 100, 150 mM, more This is best achieved by adding lactose at concentrations >300 mM. The method of any one of claims 27 or 28, wherein the lactose feed is fed to the lactose at a concentration such that a lactose concentration of at least 5 mM, preferably 10 mM or 30 mM is obtained throughout the production phase of the culture This is achieved by adding lactose to the culture. The method of any one of claims 4 to 29, wherein the cells are cultured for at least about 60, 80, 100 or about 120 hours or in a continuous manner. The method of any one of claims 4 to 30, wherein the cells are cultured in a medium comprising a carbon source, the carbon source comprising monosaccharides, disaccharides, oligosaccharides, polysaccharides, polyols, glycerol; In a complex medium of corn infusion, protein gluten, pancreas gluten or yeast extract; preferably, wherein the carbon source is selected from the list comprising the following: glucose, glycerol, fructose, sucrose, maltose, lactose, arabinose, wheat Malto-oligosaccharide, maltotriose, sorbitol, xylose, rhamnose, galactose, mannose, methanol, ethanol, trehalose, starch, cellulose, hemicellulose, molasses, corn infusion, high fruit Syrup, acetate, citrate, lactate and pyruvate. The method of any one of claims 1 to 3, 5, 11 or 12, wherein the cell-free system comprises: a. At least one alpha-1,2-fucosyltransferase that (i) comprises the polypeptide sequence of any one of SEQ ID NOs: 16 to 110, or (ii) is SEQ ID NO: 16-110 A functional homologue, variant or derivative of any one of ID NOs 16 to 110 which is the same as the alpha-1,2-rock having SEQ ID NOs 16 to 110 The full length of any of the glucosyltransferase polypeptides has at least 80% overall sequence identity and has alpha-1 for lactose and possibly any of the other receptors 2'FL and/or 3FL ,2-fucosyltransferase activity, or (iii) is a functional fragment of any one of SEQ ID NOs 16 to 110 and has alpha-1,2-fucosyltransferase activity, or (iv) Comprising a polypeptide comprising or consisting of an amino acid sequence having at least 80% sequence identity to the full-length amino acid sequence of any one of SEQ ID NOs 16 to 110 and having alpha-1,2-fucoid glycosyltransferase activity, and b. at least one α-1,3-fucosyltransferase, the fucosyltransferase (i) comprising as any one of SEQ ID NOs: 111 to 132, preferably SEQ ID NOs: 113 to any one of 132, more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NO: 113 , any one of 114, 118, 119, 120, 121 or 122, even better the polypeptide sequence of SEQ ID NO: 113 or 114, the best SEQ ID NO: 114, or (ii) the polypeptide sequence of SEQ ID NO: 111 to 114 any one of 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or Any one of 132, even better any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: The functional homologue, variant or derivative of 114, which is related to any one of SEQ ID NOs 111 to 132, preferably SEQ ID NOs: 113 to 132, more preferably Any of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NOs: 113, 114, 118, 119, 120, The full length of any one of 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: 114 of any one of the alpha-1,3-fucosyltransferase polypeptides has At least 80% overall sequence identity and alpha-1,3-fucosyltransferase activity on lactose and possibly on any of the (or) other receptors 2'FL and/or 3FL, or (iii) ) is any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128 , any one of 129, 130, 131 or 132, even more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even more preferably SEQ ID NO: 113 or 114 , the best functional fragment of SEQ ID NO: 114 with alpha-1,3-fucosyl transfer Enzymatic activity, or (iv) comprising a polypeptide comprising and any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114 , any of 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably any of SEQ ID NOs: 113, 114, 118, 119, 120, 121 or 122 or, even more preferably SEQ ID NO: 113 or 114, the full-length amino acid sequence of SEQ ID NO: 114 has or consists of an amino acid sequence having at least 80% sequence identity and has α-1,3- Fucosyltransferase activity, and c. GDP-fucose, and d. Lactose and possibly at least one of the other receptor(s). The method of claim 32, wherein the cell-free system comprises: a. at least one α-1,2-fucosyltransferase comprising as SEQ ID NOs: 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to the polypeptide sequence of any one of 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105, or (ii) is SEQ ID NO 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 A functional homologue, variant or derivative of any of the functional homologues, variants or derivatives having SEQ ID NOs 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52 , 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 of any of the α-1,2-fucosyltransferase polypeptides has at least 80% overall sequence identity over its full length and has alpha-1,2-fucosyltransferase activity on lactose and possibly on any of the other receptors 2'FL and/or 3FL(s), or (iii) is SEQ ID NO 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to A functional fragment of any one of 100, 102, 103 or 105 and having α-1,2-fucosyltransferase activity, or (iv) comprising a polypeptide comprising SEQ ID Nos 19 to 21, 23 to any of 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 A full-length amino acid sequence having or consisting of an amino acid sequence having at least 80% sequence identity and having alpha-1,2-fucosyltransferase activity, and b. at least one α-1,3-fucosyltransferase, the fucosyltransferase (i) comprising as any one of SEQ ID NOs: 111 to 132, preferably SEQ ID NOs: 113 to any one of 132, more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NO: 113 , any one of 114, 118, 119, 120, 121 or 122, even better the polypeptide sequence of SEQ ID NO: 113 or 114, the best SEQ ID NO: 114, or (ii) the polypeptide sequence of SEQ ID NO: 111 to 114 any one of 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or Any one of 132, even better any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: The functional homologue, variant or derivative of 114, which is related to any one of SEQ ID NOs 111 to 132, preferably SEQ ID NOs: 113 to 132, more preferably Any of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NOs: 113, 114, 118, 119, 120, The full length of any one of 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: 114 of any one of the alpha-1,3-fucosyltransferase polypeptides has At least 80% overall sequence identity and alpha-1,3-fucosyltransferase activity on lactose and possibly on any of the (or) other receptors 2'FL and/or 3FL, or (iii) ) is any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128 , any one of 129, 130, 131 or 132, even more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even more preferably SEQ ID NO: 113 or 114 , the best functional fragment of SEQ ID NO: 114 with alpha-1,3-fucosyl transfer Enzymatic activity, or (iv) comprising a polypeptide comprising and any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114 , any of 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably any of SEQ ID NOs: 113, 114, 118, 119, 120, 121 or 122 or, even more preferably SEQ ID NO: 113 or 114, the full-length amino acid sequence of SEQ ID NO: 114 has or consists of an amino acid sequence having at least 80% sequence identity and has α-1,3- Fucosyltransferase activity, and c. GDP-fucose, and d. Lactose and possibly at least one of the other receptor(s). The method of claim 32, wherein the cell-free system comprises: a. At least one α-1,2-fucosyltransferase, the fucosyltransferase (i) comprising as SEQ ID NO: 16 to 19 any one, preferably SEQ ID NO 16 or 19 , more preferably the polypeptide sequence of SEQ ID NO 16, or (ii) any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 or 19, more preferably a functional homologue, variation of SEQ ID NO 16 body or derivative, the functional homologue, variant or derivative with the alpha-1,2-fucoid having SEQ ID NO 16 to 19, preferably SEQ ID NO 16 or 19, more preferably SEQ ID NO 16 The full length of any of the glycosyltransferase polypeptides has at least 80% overall sequence identity and has alpha-1 for lactose and possibly any of the other receptors 2'FL and/or 3FL(s), 2-fucosyltransferase activity, or (iii) is any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 or 19, more preferably a functional fragment of SEQ ID NO 16 and having alpha-1 , 2-fucosyltransferase activity, or (iv) comprising a polypeptide comprising any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 or 19, more preferably SEQ ID NO 16 A full-length amino acid sequence having or consisting of an amino acid sequence having at least 80% sequence identity and having alpha-1,2-fucosyltransferase activity, and b. at least one α-1,3-fucosyltransferase, the fucosyltransferase (i) comprising as any one of SEQ ID NOs: 111 to 132, preferably SEQ ID NOs: 113 to any one of 132, more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NO: 113 , any one of 114, 118, 119, 120, 121 or 122, even better the polypeptide sequence of SEQ ID NO: 113 or 114, the best SEQ ID NO: 114, or (ii) the polypeptide sequence of SEQ ID NO: 111 to 114 any one of 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or Any one of 132, even better any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: The functional homologue, variant or derivative of 114, which is related to any one of SEQ ID NOs 111 to 132, preferably SEQ ID NOs: 113 to 132, more preferably Any of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NOs: 113, 114, 118, 119, 120, The full length of any one of 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: 114 of any one of the alpha-1,3-fucosyltransferase polypeptides has At least 80% overall sequence identity and alpha-1,3-fucosyltransferase activity on lactose and possibly on any of the (or) other receptors 2'FL and/or 3FL, or (iii) ) is any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128 , any one of 129, 130, 131 or 132, even more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even more preferably SEQ ID NO: 113 or 114 , the best functional fragment of SEQ ID NO: 114 with alpha-1,3-fucosyl transfer Enzymatic activity, or (iv) comprising a polypeptide comprising and any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114 , any of 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably any of SEQ ID NOs: 113, 114, 118, 119, 120, 121 or 122 or, even more preferably SEQ ID NO: 113 or 114, the full-length amino acid sequence of SEQ ID NO: 114 has or consists of an amino acid sequence having at least 80% sequence identity and has α-1,3- Fucosyltransferase activity, and c. GDP-fucose, and d. Lactose and possibly at least one of the other receptor(s). The method of any one of claims 1 to 34, wherein the separation comprises at least one of the following steps: clarification, ultrafiltration, nanofiltration, two-phase partition, reverse osmosis, microfiltration, activated carbon or carbon treatment, treatment with Nonionic surfactant treatment, enzymatic digestion, tangential flow high performance filtration, tangential flow ultrafiltration, affinity chromatography, ion exchange chromatography, hydrophobic interaction chromatography and/or gel filtration, ligand exchange layer analysis. The method of any one of claims 1 to 35, further comprising purifying any of the 2'FL, 3-FL or DiFL, preferably all of the 2'FL, 3-FL or DiFL. The method of claim 36, wherein the purification comprises at least one of the following steps: using activated charcoal or carbon, using charcoal, nanofiltration, ultrafiltration, electrophoresis, enzymatic treatment or ion exchange, using alcohol, using a hydroalcoholic mixture, Crystallization, evaporation, precipitation, drying, spray drying, freeze drying, spray freeze drying, freeze spray drying, band drying, belt drying, vacuum belt drying, vacuum belt drying, Drum drying, roller drying, vacuum drum drying or vacuum roller drying. The method of any one of claims 1 to 37, wherein the DiFL is at least 80 as measured by the total amount of 2'FL, 3-FL and DiFL produced in the 2'FL, 3-FL and DiFL mixture % purity was purified from the 2'FL, 3-FL and DiFL mixture. A metabolically engineered cell for the production of DiFL or a mixture of DiFL, 2'FL and/or 3-FL, wherein the cell: a. Represents at least two fucosyltransferases capable of transferring fucose residues from a GDP-fucose donor to an acceptor lactose and an acceptor 2'FL and/or any one or more of 3-FL, and b. capable of synthesizing GDP-fucose, wherein the GDP-fucose is the donor of these fucosyltransferases, c. Preferably wherein the cell is metabolically engineered for the production of DiFL or a mixture of diFL, 2'FL and/or 3-FL. The cell of claim 39, wherein the cell is modified with at least one gene expression module, characterized in that the expression from any of these expression modules is persistent or produced by a natural inducer, preferably persistent. The cell of claim 40, wherein the cell comprises multiple copies of the same coding DNA sequence encoding a protein. The cell of any one of claims 30 and 31, wherein the cell exhibits: a. At least one alpha-1,2-fucosyltransferase that (i) comprises the polypeptide sequence of any one of SEQ ID NOs: 16 to 110, or (ii) is SEQ ID NO: 16-110 A functional homologue, variant or derivative of any one of ID NOs 16 to 110 which is the same as the alpha-1,2-rock having SEQ ID NOs 16 to 110 The full length of any of the glucosyltransferase polypeptides has at least 80% overall sequence identity and has alpha-1 for lactose and possibly any of the other receptors 2'FL and/or 3FL ,2-fucosyltransferase activity, or (iii) is a functional fragment of any one of SEQ ID NOs 16 to 110 and has alpha-1,2-fucosyltransferase activity, or (iv) Comprising a polypeptide comprising or consisting of an amino acid sequence having at least 80% sequence identity to the full-length amino acid sequence of any one of SEQ ID NOs 16 to 110 and having alpha-1,2-fucoid glycosyltransferase activity, and b. at least one α-1,3-fucosyltransferase, the fucosyltransferase (i) comprising as any one of SEQ ID NOs: 111 to 132, preferably SEQ ID NOs: 113 to any one of 132, more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NO: 113 , any one of 114, 118, 119, 120, 121 or 122, even better the polypeptide sequence of SEQ ID NO: 113 or 114, the best SEQ ID NO: 114, or (ii) the polypeptide sequence of SEQ ID NO: 111 to 114 any one of 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or Any one of 132, even better any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: The functional homologue, variant or derivative of 114, which is related to any one of SEQ ID NOs 111 to 132, preferably SEQ ID NOs: 113 to 132, more preferably Any of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NOs: 113, 114, 118, 119, 120, The full length of any one of 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: 114 of any one of the alpha-1,3-fucosyltransferase polypeptides has At least 80% overall sequence identity and alpha-1,3-fucosyltransferase activity on lactose and possibly on any of the (or) other receptors 2'FL and/or 3FL, or (iii) ) is any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128 , any one of 129, 130, 131 or 132, even more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even more preferably SEQ ID NO: 113 or 114 , the best functional fragment of SEQ ID NO: 114 with alpha-1,3-fucosyl transfer Enzymatic activity, or (iv) comprising a polypeptide comprising and any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114 , any of 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably any of SEQ ID NOs: 113, 114, 118, 119, 120, 121 or 122 or, even more preferably SEQ ID NO: 113 or 114, the full-length amino acid sequence of SEQ ID NO: 114 has or consists of an amino acid sequence having at least 80% sequence identity and has α-1,3- Fucosyltransferase activity. The cell of claim 42, wherein the cell exhibits: a. at least one α-1,2-fucosyltransferase comprising as SEQ ID NOs: 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to the polypeptide sequence of any one of 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105, or (ii) is SEQ ID NO 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 A functional homologue, variant or derivative of any of the functional homologues, variants or derivatives having SEQ ID NOs 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52 , 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 of any of the α-1,2-fucosyltransferase polypeptides has at least 80% overall sequence identity over its full length and has alpha-1,2-fucosyltransferase activity on lactose and possibly on any of the other receptors 2'FL and/or 3FL(s), or (iii) is SEQ ID NO 19 to 21, 23 to 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to A functional fragment of any one of 100, 102, 103 or 105 and having α-1,2-fucosyltransferase activity, or (iv) comprising a polypeptide comprising and SEQ ID NOs 19 to 21, 23 to any of 38, 40 to 44, 46 to 48, 50 to 52, 54 to 61, 64, 65, 69 to 73, 76 to 86, 88 to 91, 96 to 100, 102, 103 or 105 A full-length amino acid sequence having or consisting of an amino acid sequence having at least 80% sequence identity and having alpha-1,2-fucosyltransferase activity, and b. at least one α-1,3-fucosyltransferase, the fucosyltransferase (i) comprising as any one of SEQ ID NOs: 111 to 132, preferably SEQ ID NOs: 113 to any one of 132, more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NO: 113 , any one of 114, 118, 119, 120, 121 or 122, even better the polypeptide sequence of SEQ ID NO: 113 or 114, the best SEQ ID NO: 114, or (ii) the polypeptide sequence of SEQ ID NO: 111 to 114 any one of 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or Any one of 132, even better any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: The functional homologue, variant or derivative of 114, which is related to any one of SEQ ID NOs 111 to 132, preferably SEQ ID NOs: 113 to 132, more preferably Any of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NOs: 113, 114, 118, 119, 120, The full length of any one of 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: 114 of any one of the alpha-1,3-fucosyltransferase polypeptides has At least 80% overall sequence identity and alpha-1,3-fucosyltransferase activity on lactose and possibly on any of the (or) other receptors 2'FL and/or 3FL, or (iii) ) is any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128 , any one of 129, 130, 131 or 132, even more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even more preferably SEQ ID NO: 113 or 114 , the best functional fragment of SEQ ID NO: 114 with alpha-1,3-fucosyl transfer Enzymatic activity, or (iv) comprising a polypeptide comprising and any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114 , any of 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably any of SEQ ID NOs: 113, 114, 118, 119, 120, 121 or 122 or, even more preferably SEQ ID NO: 113 or 114, the full-length amino acid sequence of SEQ ID NO: 114 has or consists of an amino acid sequence having at least 80% sequence identity and has α-1,3- Fucosyltransferase activity. The cell of claim 42, wherein the cell exhibits: a. At least one α-1,2-fucosyltransferase, the fucosyltransferase (i) comprising as SEQ ID NO: 16 to 19 any one, preferably SEQ ID NO 16 or 19 , more preferably the polypeptide sequence of SEQ ID NO 16, or (ii) any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 or 19, more preferably a functional homologue, variation of SEQ ID NO 16 body or derivative, the functional homologue, variant or derivative with the alpha-1,2-fucoid having SEQ ID NO 16 to 19, preferably SEQ ID NO 16 or 19, more preferably SEQ ID NO 16 The full length of any of the glycosyltransferase polypeptides has at least 80% overall sequence identity and has alpha-1 for lactose and possibly any of the other receptors 2'FL and/or 3FL(s), 2-fucosyltransferase activity, or (iii) is any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 or 19, more preferably a functional fragment of SEQ ID NO 16 and having alpha-1 , 2-fucosyltransferase activity, or (iv) comprising a polypeptide comprising any one of SEQ ID NOs 16 to 19, preferably SEQ ID NO 16 or 19, more preferably SEQ ID NO 16 A full-length amino acid sequence having or consisting of an amino acid sequence having at least 80% sequence identity and having alpha-1,2-fucosyltransferase activity, and b. at least one α-1,3-fucosyltransferase, the fucosyltransferase (i) comprising as any one of SEQ ID NOs: 111 to 132, preferably SEQ ID NOs: 113 to any one of 132, more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NO: 113 , any one of 114, 118, 119, 120, 121 or 122, even better the polypeptide sequence of SEQ ID NO: 113 or 114, the best SEQ ID NO: 114, or (ii) the polypeptide sequence of SEQ ID NO: 111 to 114 any one of 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or Any one of 132, even better any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: The functional homologue, variant or derivative of 114, which is related to any one of SEQ ID NOs 111 to 132, preferably SEQ ID NOs: 113 to 132, more preferably Any of SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even better SEQ ID NOs: 113, 114, 118, 119, 120, The full length of any one of 121 or 122, even better SEQ ID NO: 113 or 114, best SEQ ID NO: 114 of any one of the alpha-1,3-fucosyltransferase polypeptides has At least 80% overall sequence identity and alpha-1,3-fucosyltransferase activity on lactose and possibly on any of the (or) other receptors 2'FL and/or 3FL, or (iii) ) is any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114, 118, 119, 120, 121, 122, 128 , any one of 129, 130, 131 or 132, even more preferably any one of SEQ ID NO: 113, 114, 118, 119, 120, 121 or 122, even more preferably SEQ ID NO: 113 or 114 , the best functional fragment of SEQ ID NO: 114 with alpha-1,3-fucosyl transfer Enzymatic activity, or (iv) comprising a polypeptide comprising and any one of SEQ ID NOs 111 to 132, preferably any one of SEQ ID NOs: 113 to 132, more preferably SEQ ID NOs: 113, 114 , any of 118, 119, 120, 121, 122, 128, 129, 130, 131 or 132, even more preferably any of SEQ ID NOs: 113, 114, 118, 119, 120, 121 or 122 or, even more preferably SEQ ID NO: 113 or 114, the full-length amino acid sequence of SEQ ID NO: 114 has or consists of an amino acid sequence having at least 80% sequence identity and has α-1,3- Fucosyltransferase activity. The cell of any one of claims 39 to 44, wherein the cell produces lactose for the synthesis of any of the 2'FL, 3-FL and DiFL, preferably the cell further produces the one or more other receptors at least one of the bodies. The cell of any one of claims 39 to 45, wherein the cell uses one or more precursors for the synthesis of any of the 2'FL, 3-FL and DiFL. The cell of any one of claims 39 to 46, wherein the precursor(s) and/or receptor are fully converted to any of the 2'FL, 3-FL and DiFL. The cell of any one of claims 39 to 47, wherein the cell is modified in the expression or activity of at least one of the fucosyltransferases. The cell of any one of claims 39 to 48, wherein the cell is modified to produce GDP-fucose. The cell of any one of claims 39 to 49, wherein the cell is modified for enhanced GDP-fucose production compared to an unmodified precursor cell, and wherein the modification is selected from the group consisting of Group: UDP-glucose: Deletion of the gene encoding undecyl isopentenyl-phosphoglucose-1-phosphotransferase, overexpression of the gene encoding GDP-L-fucose synthase, GDP-mannose 4,6-dehydration Overexpression of gene encoding enzyme, overexpression of gene encoding mannose-1-phosphate guanosyltransferase, overexpression of gene encoding phosphomannose mutase, or overexpression of gene encoding mannose-6-phosphate isomerase Performance. The cell of any one of claims 39 to 50, wherein the cell is resistant to lactose killing when grown in an environment where lactose is combined with one or more other carbon sources. The cell of any one of claims 39 to 51, wherein the cell intracellularly produces the DiFL or a mixture of the DiFL, 2'FL and/or 3-FL and wherein the produced DiFL, 2'FL and/or 3-FL - Some or substantially all of the FL remains intracellular and/or is excreted outside the cell via passive or active transport. The cell of any one of claims 39 to 52, wherein the cell is further genetically modified for i) Modified expression of endogenous membrane proteins, and/or ii) the modified activity of the endogenous membrane protein, and/or iii) Representation of homologous membrane proteins, and/or iv) expression of heterologous membrane proteins, wherein the membrane protein participates in the secretion of any one of the 2'FL, 3-FL and DiFL from the mixture to the outside of the cell, preferably, the membrane protein participates in the secretion of the 2'FL from the mixture to the outside of the cell All of 2'FL, 3-FL and DiFL. The cell of any one of claims 39 to 53, wherein the cell is further genetically modified for i) Modified expression of endogenous membrane proteins, and/or ii) the modified activity of the endogenous membrane protein, and/or iii) Representation of homologous membrane proteins, and/or iv) expression of heterologous membrane proteins, wherein the membrane protein is involved in the uptake of lactose and/or at least one other receptor 2'FL and/or 3FL for the synthesis of any of the 2'FL, 3-FL and DiFL. The cell of any one of claims 53 or 54, wherein the membrane protein is selected from the list comprising transport proteins, P-P-bond hydrolysis-driven transport proteins, β-barrel porins, accessory transport proteins, Putative transporter and phosphotransfer driven group translocation protein, Preferably, these transport proteins comprise MFS transport proteins, carbohydrate efflux transport proteins and chelating ferritin export proteins, Preferably, the P-P-bond hydrolysis-driven transport proteins comprise ABC transport proteins and chelatin export proteins. The cell of any one of claims 53 to 55, wherein the membrane protein improves the production of any one of the 2'FL, 3-FL and DiFL and/or enables and/or enhances the 2'FL, 3-FL and DiFL - Outflow of either FL and DiFL. The cell of any one of claims 39 to 56 or the method of any one of claims 4 to 31 and 35 to 38, wherein the cell is a bacterium, fungus, yeast, plant cell, animal cell or protozoal cell , - preferably, the bacteria are Escherichia coli ( Escherichia coli ) strains, more preferably Escherichia coli strains of K-12 strains, even more preferably the Escherichia coli K-12 strains are Escherichia coli MG1655, - preferably, the The fungus belongs to a genus selected from the group comprising Rhizopus , Dictyostelium , Penicillium , Mucor or Aspergillus , - more Preferably, the yeast belongs to a genus selected from the group comprising: Saccharomyces , Zygosaccharomyces , Pichia , Komagataella, Hansenula Genus Hansenula , Yarrowia , Starmerella , Kluyveromyces or Debaromyces , - preferably, the plant cell is an algae cells or derived from tobacco, alfalfa, rice, tomato, cotton, rapeseed, soybean, maize or corn plants, - preferably, the animal cells are derived from non-human mammals, birds, fish, invertebrates, reptiles, Amphibians or insects, or genetically modified cell lines derived from human cells excluding embryonic stem cells, more preferably the human and non-human mammalian cells are epithelial cells, embryonic kidney cells, fibroblasts, COS cells, Chinese Hamster ovary (CHO) cells, murine myeloma cells, NIH-3T3 cells, non-mammary adult stem cells or derivatives thereof, preferably the insect cells are derived from Spodoptera frugiperda , Bombyx mori , Mamestra brassicae , Trichoplusia ni or Drosophila melanogaster , - preferably, the protozoal cell is a Leishmania tarentolae cell. The cell of claim 57 or the method of claim 57, wherein the cell is a viable Gram-negative bacterium comprising a reduced or eliminated synthesis of poly-N compared to an unmodified precursor cell -Acetyl-glucosamine (PNAG), Enterobacterial common antigen (ECA), cellulose, colanic acid, core oligosaccharide, osmotic periplasmic glucan (OPG), glycerol glucoside, Glycans and/or trehalose. Use of a cell according to any one of claims 39 to 58 or a method according to any one of claims 1 to 38, 57 and 58 for the production of a mixture of DiFL, 2'FL and/or 3-FL . A method for producing 2',3-difucosyllactose (DiFL) comprising the steps of: i) provide a cell as in any one of claims 39 to 58, and, ii) culturing the cells under conditions that allow expression of the fucosyltransferases and synthesis of the GDP-fucose, iii) Preferably, at least one of the 2'FL, 3-FL or DiFL is isolated from the culture, more preferably DiFL is isolated from the culture, even more preferably the 2'FL, 3-FL is isolated from the culture - All of FL and DiFL. A use of a cell as claimed in any one of claims 39 to 58 for the production of DiFL.