WO2019096903A1

WO2019096903A1 - New galactanases (ec 3.2.1.89) for use in soy processing

Info

Publication number: WO2019096903A1
Application number: PCT/EP2018/081367
Authority: WO
Inventors: Laerke Tvedebrink HAAHR; Lone CARSTENSEN; Diego Filipe ROSSO
Original assignee: Novozymes A/S
Priority date: 2017-11-20
Filing date: 2018-11-15
Publication date: 2019-05-23

Abstract

The present invention relates to methods, uses and animal feeds employing polypeptides having galactanase activity, wherein the galactanases are used for processing soy-based material.

Description

NEW GALACTANASES (EC 3.2.1.89) FOR USE IN SOY PROCESSING

Reference to sequence listing

This application contains a Sequence Listing in computer readable form. The computer reada- ble form is incorporated herein by reference.

INDEX TO SEQUENCE LISTING

SEQ ID NO: 1 is the cDNA sequence of the GH53 galactanase as isolated from Cohnella sp- 60555.

SEQ ID NO: 2 is the amino acid sequence as deduced from SEQ ID NO: 1.

SEQ ID NO: 3 is the amino acid sequence of the mature GH53 galactanase from Cohnella sp- 60555.

SEQ ID NO: 4 is the amino acid sequence of SEQ ID NO: 3 with His-tag.

SEQ ID NO: 5 is the cDNA sequence of the GH53 galactanase as isolated from Cohnella xylani- lytica.

SEQ ID NO: 6 is the amino acid sequence as deduced from SEQ ID NO: 5.

SEQ ID NO: 7 is the amino acid sequence of the mature GH53 galactanase from Cohnella xy- lanilytica.

SEQ ID NO: 8 is the amino acid sequence of SEQ ID NO: 7 with His-tag.

SEQ ID NO: 9 is the cDNA sequence of the GH53 galactanase as isolated from Paenibacillus sp-18179.

SEQ ID NO: 10 is the amino acid sequence as deduced from SEQ ID NO: 9.

SEQ ID NO: 11 is the amino acid sequence of the mature GH53 galactanase from Paenibacillus sp-18179.

SEQ ID NO: 12 is the amino acid sequence of SEQ ID NO: 11 with His-tag.

SEQ ID NO: 13 is the cDNA sequence of the GH53 galactanase as isolated from Paenibacillus peoriae.

SEQ ID NO: 14 is the amino acid sequence as deduced from SEQ ID NO: 13.

SEQ ID NO: 15 is the amino acid sequence of the mature GH53 galactanase from Paenibacillus peoriae.

SEQ ID NO: 16 is the amino acid sequence of SEQ ID NO: 15 with His-tag. SEQ ID NO: 17 is the cDNA sequence of the GH53 galactanase as isolated from Paenibacillus xylanexedens.

SEQ ID NO: 18 is the amino acid sequence as deduced from SEQ ID NO: 17.

SEQ ID NO: 19 is the amino acid sequence of the mature GH53 galactanase from Paenibacillus xylanexedens.

SEQ ID NO: 20 is the amino acid sequence of SEQ ID NO: 19 with His-tag.

SEQ ID NO: 21 is the cDNA sequence of the GH53 galactanase as isolated from Cohnella laeviribosi.

SEQ ID NO: 22 is the amino acid sequence as deduced from SEQ ID NO: 21.

SEQ ID NO: 23 is the amino acid sequence of the mature GH53 galactanase from Cohnella laeviribosi.

SEQ ID NO: 24 is the amino acid sequence of SEQ ID NO: 23 with His-tag.

SEQ ID NO: 25 is the amino acid sequence of the mature GH53 galactanase from Humicola insolens as disclosed in WO1997/032014.

SEQ ID NO: 26 is the conserved motif GV[T/M]PD[W/M]VQ[I/V]GNE.

SEQ ID NO: 27 is the conserved motif WADP[A/G]xQxKPxAW.

SEQ ID NO: 28 is the Bacillus clausii secretion signal.

SEQ ID NO: 29 is the cDNA sequence of the GH53 galactanase as isolated from Xanthomonas cynarae.

SEQ ID NO: 30 is the amino acid sequence as deduced from SEQ ID NO: 29.

SEQ ID NO: 31 is the amino acid sequence of the mature GH53 galactanase from Xanthomo nas cynarae.

SEQ ID NO: 32 is the amino acid sequence of SEQ ID NO: 31 with His-tag.

FIELD OF THE INVENTION

The present invention relates to compositions comprising polypeptides having galactanase ac- tivity for use in soy processing. The present invention further relates to polypeptides having ga- lactanase activity and polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as meth- ods of producing and using the polypeptides. BACKGROUND OF THE INVENTION

Soybean is a species of legume native to East Asia and is the second biggest crop globally and the biggest protein source. Soy bean can be manufactured to produce e.g. soybean meal (SBM), soy protein concentrate (SPC) and fermented soybean meal (FSBM).

Production of soy bean meal

A schematic view of typical soybean processing is shown in Fig. 1. Preferably the first step is selection and cleaning of high quality soybeans. The soybeans are typically dehulled, cracked, flaked and steam conditioned before they enter a solvent extraction process. The oil is typically removed and further processed. The crude soy oil can be fractionated and refined to soy oil and lecithin. The soy oil can be used for various purposes e.g. in the food industry. After extraction, defatted flakes remain which can be used in various further processing steps. The most com- monly known and most widely used product that is made from defatted flakes is soybean meal. The flakes are typically steam toasted to remove the residual solvent and to inactivate the heat labile anti-nutritional factors (ANFs). Depending on the amount of soy hulls that are added back to the flakes, the protein level of soybean meal is adjusted. The major application of soybean meal is in animal nutrition.

Production of soy protein concentrate (SPC)

In order to avoid browning reactions in the production of specialty soy proteins, the defatted flakes do not enter the common desolventizer/toaster. Instead, the hexane is removed without altering the functional characteristics of the soybean proteins by low heat vacuum drying. This yields the so-called "white flakes". The removal of the soluble carbohydrates from the white flakes leads to SPC. The vast majority of SPC is produced by an extraction method. Aqueous alcohol leaching selectively removes the soluble carbohydrates, while keeping most of the in- soluble protein. With this processing step other ANFs than the oligosaccharides are simultane- ously removed- such as e.g. estrogens and antigenic factors. The SPC is distinctly different to the soybean meal as it contains just traces of oligosaccharides and the antigenic substances glycinin and b-conglycinin. Therefore it can be used e.g. in milk replacer feed for non-ruminant calves, piglet pre-starter feed, pet food, feed for fur bearing animals and in aquaculture

Production of fermented soybean meal (FSBM)

Soybean meal (SBM) is the major source of protein in swine diets used in several countries. The amino acid profile of SBM fits well with that of corn and some other cereal grains to balance swine diets; however, its inclusion in diets for newly weaned pigs is limited due to some anti- nutritional factors and antigenic soybean proteins causing hypersensitivity. Detrimental effects due to SBM in young pigs include reduction in growth rate, nutrient digestibility and intestinal villus height, and a greater immune response to soybean proteins. Fermentation has proven to improve the nutritional value of soybean by increasing the bioavailability of nutrients and reduc- ing anti-nutritional factors. The fermentation process can be used as a strategy to reduce the antigenicity of soybean proteins and consequently avoid detrimental effects in weaned pigs. However, the efficiency of fermentation to reduce soybean proteins immunoreactivity depends on many factors such as starter culture, type of fermentation, fermentation time, and raw mate- rial particle size. Most FSBM is produced by solid state fermentation with a mix of bacterial and fungal strains e.g. bacillus, lactobacillus and saccharomyces cerevisae. Enzymes such as pro- teases and carbohydrate active enzymes can be included in the fermentation process to achieve a better final product, by breaking down antigenic proteins, and releasing more sugars for the fermentation organisms to use.

Galactanase used in soy processing solubilizes sugars from soybean meal. In the SPC (soy protein concentrate) process, this means that more sugars are washed away during the ethanol wash, leading to a higher protein concentration in the final product. In FSBM (Fermented soy- bean meal), the sugars solubilized by galactanase are converted to lactic acid and CO₂ by mi- croorganisms leading to a final product with more lactic acid and higher protein concentration due to CO₂ evaporation. Compared to other commercial enzymes, galactanase releases signifi cantly more soluble sugars from soybean meal.

WO 01/59083 suggests the use of bacterial galactanase, e.g. from Bacillus pumilus, for modifying and/or improving the nutritional value of an animal feed.

The galactanases of the present invention have high activity on soybean meal and are e.g. higher compared to the benchmark galactanases from H. insolens and B. licheniformis.

SUMMARY OF THE INVENTION

The invention relates to a method for modifying and/or improving the nutritional value of a soy- based material, the method comprising the steps of i) providing a soy-based material and ii) treating the soy-based material with a polypeptide having galactanase activity. The present application further relates to isolated polypeptides having galactanase activity; compositions comprising polypeptides having galactanase activity; animal feed or animal additives comprising the polypeptide(s) of the invention and use of the polypeptide(s); polynucleotides encoding the polypeptides of the present invention; nucleic acid constructs; expression vectors; recombinant host cells comprising the polynucleotides; methods of producing the polypeptides; and compositions thereof. The present invention further relates to compositions comprising one or more GH53 polypeptides having galactanase activity. BRIEF DESCRIPTION OF DRAWINGS

Fig. 1 shows the typical steps in manufacture of soy products.

DEFINITIONS

Allelic variant: The term“allelic variant” means any of two or more alternative forms of a gene occupying the same chromosomal locus. Allelic variation arises naturally through mutation, and may result in polymorphism within populations. Gene mutations can be silent (no change in the encoded polypeptide) or may encode polypeptides having altered amino acid sequences. An allelic variant of a polypeptide is a polypeptide encoded by an allelic variant of a gene.

Animal: The term“animal” refers to all animals except humans. Examples of animals are non- ruminants, and ruminants. Ruminant animals include, for example, animals such as sheep, goats, cattle, e.g. beef cattle, cows, and young calves, deer, yank, camel, llama and kangaroo. Non-ruminant animals include mono-gastric animals, e.g. pigs or swine (including, but not lim ited to, piglets, growing pigs, and sows); poultry such as turkeys, ducks and chicken (including but not limited to broiler chicks, layers); horses (including but not limited to hotbloods, cold- bloods and warm bloods), young calves; fish (including but not limited to amberjack, arapaima, barb, bass, bluefish, bocachico, bream, bullhead, cachama, carp, catfish, catla, chanos, char, cichlid, cobia, cod, crappie, dorada, drum, eel, goby, goldfish, gourami, grouper, guapote, hali but, java, labeo, lai, loach, mackerel, milkfish, mojarra, mudfish, mullet, paco, pearlspot, pejer- rey, perch, pike, pompano, roach, salmon, sampa, sauger, sea bass, seabream, shiner, sleeper, snakehead, snapper, snook, sole, spinefoot, sturgeon, sunfish, sweetfish, tench, terror, tilapia, trout, tuna, turbot, vendace, walleye and whitefish); and crustaceans (including but not limited to shrimps and prawns).

Animal feed: The term“animal feed” refers to any compound, preparation, or mixture suitable for, or intended for intake by an animal. Animal feed for a mono-gastric animal typically corn- prises concentrates as well as vitamins, minerals, enzymes, direct fed microbial, amino acids and/or other feed ingredients (such as in a premix) whereas animal feed for ruminants generally comprises forage (including roughage and silage) and may further comprise concentrates as well as vitamins, minerals, enzymes direct fed microbial, amino acid and/or other feed ingredi- ents (such as in a premix).

Body Weight Gain: The term“body weight gain” means an increase in live weight of an animal during a given period of time e.g. the increase in weight from day 1 to day 21.

cDNA: The term "cDNA" means a DNA molecule that can be prepared by reverse transcription from a mature, spliced, mRNA molecule obtained from a eukaryotic or prokaryotic cell. cDNA lacks intron sequences that may be present in the corresponding genomic DNA. The initial, pri- mary RNA transcript is a precursor to mRNA that is processed through a series of steps, includ ing splicing, before appearing as mature spliced mRNA. Coding sequence: The term“coding sequence” means a polynucleotide, which directly speci- fies the amino acid sequence of a polypeptide. The boundaries of the coding sequence are generally determined by an open reading frame, which begins with a start codon such as ATG, GTG, or TTG and ends with a stop codon such as TAA, TAG, or TGA. The coding sequence may be a genomic DNA, cDNA, synthetic DNA, or a combination thereof.

Control sequences: The term“control sequences” means nucleic acid sequences necessary for expression of a polynucleotide encoding a mature polypeptide of the present invention. Each control sequence may be native (/^'.e., from the same gene) or foreign (/^'.e., from a different gene) to the polynucleotide encoding the polypeptide or native or foreign to each other. Such control sequences include, but are not limited to, a leader, polyadenylation sequence, propeptide se- quence, promoter, signal peptide sequence, and transcription terminator. At a minimum, the control sequences include a promoter, and transcriptional and translational stop signals. The control sequences may be provided with linkers for the purpose of introducing specific restriction sites facilitating ligation of the control sequences with the coding region of the polynucleotide encoding a polypeptide.

Expression: The term“expression” includes any step involved in the production of a polypep- tide including, but not limited to, transcription, post-transcriptional modification, translation, post- translational modification, and secretion.

Expression vector: The term“expression vector” means a linear or circular DNA molecule that comprises a polynucleotide encoding a polypeptide and is operably linked to control sequences that provide for its expression.

Feed Conversion Ratio: The term“feed conversion ratio” the amount of feed fed to an animal to increase the weight of the animal by a specified amount. An improved feed conversion ratio means a lower feed conversion ratio. By "lower feed conversion ratio" or "improved feed con- version ratio" it is meant that the use of a feed additive composition in feed results in a lower amount of feed being required to be fed to an animal to increase the weight of the animal by a specified amount compared to the amount of feed required to increase the weight of the animal by the same amount when the feed does not comprise said feed additive composition.

Feed efficiency: The term“feed efficiency” means the amount of weight gain per unit of feed when the animal is fed ad-libitum or a specified amount of food during a period of time. By "in- creased feed efficiency" it is meant that the use of a feed additive composition according the present invention in feed results in an increased weight gain per unit of feed intake compared with an animal fed without said feed additive composition being present.

Fragment: The term“fragment” means a polypeptide having one or more (e.g., several) amino acids absent from the amino and/or carboxyl terminus of a mature polypeptide or domain; wherein the fragment has galactanase activity. In another embodiment, the fragment comprises at least 90% of the length of the mature poly- peptide, such as at least 284 amino acids of SEQ ID NO: 3, at least 286 amino acids of SEQ ID NO: 7, at least 464 amino acids of SEQ ID NO: 11 , at least 285 amino acids of SEQ ID NO: 15, at least 284 amino acids of SEQ ID NO: 19, at least 284 amino acids of SEQ ID NO: 23 or at least 276 amino acids of SEQ ID NO: 31.

In another embodiment, the fragment comprises at least 92% of the length of the mature poly- peptide, such as at least 290 amino acids of SEQ ID NO: 3, at least 292 amino acids of SEQ ID NO: 7, at least 474 amino acids of SEQ ID NO: 11 , at least 291 amino acids of SEQ ID NO: 15, at least 290 amino acids of SEQ ID NO: 19, at least 290 amino acids of SEQ ID NO: 23 or at least 282 amino acids of SEQ ID NO: 31.

In another embodiment, the fragment comprises at least 94% of the length of the mature poly- peptide, such as at least 297 amino acids of SEQ ID NO: 3, at least 298 amino acids of SEQ ID NO: 7, at least 485 amino acids of SEQ ID NO: 11 , at least 297 amino acids of SEQ ID NO: 15, at least 297 amino acids of SEQ ID NO: 19, at least 297 amino acids of SEQ ID NO: 23 or at least 288 amino acids of SEQ ID NO: 31.

In another embodiment, the fragment comprises at least 96% of the length of the mature poly- peptide, such as at least 303 amino acids of SEQ ID NO: 3, at least 305 amino acids of SEQ ID NO: 7, at least 495 amino acids of SEQ ID NO: 11 , at least 304 amino acids of SEQ ID NO: 15, at least 303 amino acids of SEQ ID NO: 19, at least 303 amino acids of SEQ ID NO: 23 or at least 295 amino acids of SEQ ID NO: 31.

In another embodiment, the fragment comprises at least 98% of the length of the mature poly- peptide, such as at least 309 amino acids of SEQ ID NO: 3, at least 31 1 amino acids of SEQ ID NO: 7, at least 505 amino acids of SEQ ID NO: 11 , at least 310 amino acids of SEQ ID NO: 15, at least 309 amino acids of SEQ ID NO: 19, at least 309 amino acids of SEQ ID NO: 23 or at least 301 amino acids of SEQ ID NO: 31.

In another embodiment, the fragment comprises at least 99% of the length of the mature poly- peptide, such as at least 312 amino acids of SEQ ID NO: 3, at least 314 amino acids of SEQ ID NO: 7, at least 510 amino acids of SEQ ID NO: 11 , at least 313 amino acids of SEQ ID NO: 15, at least 312 amino acids of SEQ ID NO: 19, at least 312 amino acids of SEQ ID NO: 23 or at least 304 amino acids of SEQ ID NO: 31.

Galactanase: The term“galactanase”, also called endo-1 ,4-3-galactanase, means an arabino- galactan endo-b-1 ,4-galactanase (E.C. 3.2.1.89) that catalyses the hydrolysis of (1 4)-3-D- galactosidic linkages in type I arabinogalactans. Galactanase activity can be determined by re- ducing ends using the colorimetric assay developed by Lever (Analytical Biochemistry 47, 273- 279, 1972). The galactanase produces reducing end sugars which react with PAHBAH generat- ing an increase of colour which is proportional to the enzyme activity under the conditions used in the assay. A detailed assay can be found in the galactanase assay as described herein.

The galactanases of the present invention have at least 60%, such as at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95% or at least 100% of the galactanase activity of the polypeptide of SEQ ID NO: 3.

Host cell: The term "host cell" means any cell type that is susceptible to transformation, trans- fection, transduction, or the like with a nucleic acid construct or expression vector comprising a polynucleotide of the present invention. The term“host cell” encompasses any progeny of a parent cell that is not identical to the parent cell due to mutations that occur during replication.

Improve nutritional value: the term“improved nutritional value” means that the soy-based ma- terial treated with one or more of the galactanases of the invention has a higher nutritional value compared to similar soy-based material that has not been treated with one or more galac- tanases (termed“untreated material”). The soy-based material with improved nutritional value can have a higher protein content than the untreated material e.g. such that the protein content is increased by at least 1 %, such as at least 2%, such as at least 3%, such as at least 4%, such as at least 5%, such as at least 6%, such as at least 7%, such as at least 8%, such as at least 9% or such as at least 10%. The soy-based material with improved nutritional value can have an increased NDF digestibility than the untreated material e.g. such that the NDF digestibility is increased by at least 1%, such as at least 2%, such as at least 3%, such as at least 4%, such as at least 5%, such as at least 6%, such as at least 7%, such as at least 8%, such as at least 9% or such as at least 10%.“Improved nutritional value” can mean improved“Nutrient Digesti- bility” of the soy-based material treated with one or more of the galactanases of the invention.

Isolated: The term“isolated” means a substance in a form or environment that does not occur in nature. Non-limiting examples of isolated substances include (1 ) any non-naturally occurring substance, (2) any substance including, but not limited to, any enzyme, variant, nucleic acid, protein, peptide or cofactor, that is at least partially removed from one or more or all of the natu- rally occurring constituents with which it is associated in nature; (3) any substance modified by the hand of man relative to that substance found in nature; or (4) any substance modified by increasing the amount of the substance relative to other components with which it is naturally associated (e.g., recombinant production in a host cell; multiple copies of a gene encoding the substance; and use of a stronger promoter than the promoter naturally associated with the gene encoding the substance).

Mature polypeptide: The term“mature polypeptide” means a polypeptide in its final form fol lowing translation and any post-translational modifications, such as N-terminal processing, C-terminal truncation, glycosylation, phosphorylation, etc. In one embodiment, the mature polypeptide is amino acids 1 to 316 of SEQ ID NO: 2 and amino acids -32 to -1 of SEQ ID NO: 2 are a signal peptide. In another embodiment, the mature poly- peptide is amino acids 1 to 316 of SEQ ID NO: 3. In an alternative embodiment, the mature pol- ypeptide is amino acids 1 to 324 of SEQ ID NO: 4.

In one embodiment, the mature polypeptide is amino acids 1 to 318 of SEQ ID NO: 6 and amino acids -29 to -1 of SEQ ID NO: 6 are a signal peptide. In another embodiment, the mature poly- peptide is amino acids 1 to 318 of SEQ ID NO: 7. In an alternative embodiment, the mature pol- ypeptide is amino acids 1 to 326 of SEQ ID NO: 8.

In one embodiment, the mature polypeptide is amino acids 1 to 516 of SEQ ID NO: 10 and ami- no acids -29 to -1 of SEQ ID NO: 10 are a signal peptide. In another embodiment, the mature polypeptide is amino acids 1 to 516 of SEQ ID NO: 1 1. In an alternative embodiment, the ma- ture polypeptide is amino acids 1 to 524 of SEQ ID NO: 12.

In one embodiment, the mature polypeptide is amino acids 1 to 317 of SEQ ID NO: 14 and ami- no acids -33 to -1 of SEQ ID NO: 14 are a signal peptide. In another embodiment, the mature polypeptide is amino acids 1 to 317 of SEQ ID NO: 15. In an alternative embodiment, the ma- ture polypeptide is amino acids 1 to 325 of SEQ ID NO: 16.

In one embodiment, the mature polypeptide is amino acids 1 to 316 of SEQ ID NO: 18 and ami- no acids -33 to -1 of SEQ ID NO: 18 are a signal peptide. In another embodiment, the mature polypeptide is amino acids 1 to 316 of SEQ ID NO: 19. In an alternative embodiment, the ma- ture polypeptide is amino acids 1 to 324 of SEQ ID NO: 20.

In one embodiment, the mature polypeptide is amino acids 1 to 316 of SEQ ID NO: 22 and ami- no acids -31 to -1 of SEQ ID NO: 22 are a signal peptide. In another embodiment, the mature polypeptide is amino acids 1 to 316 of SEQ ID NO: 23. In an alternative embodiment, the ma- ture polypeptide is amino acids 1 to 324 of SEQ ID NO: 24.

In one embodiment, the mature polypeptide is amino acids 1 to 307 of SEQ ID NO: 30 and ami- no acids -26 to -1 of SEQ ID NO: 30 are a signal peptide. In another embodiment, the mature polypeptide is amino acids 1 to 307 of SEQ ID NO: 31. In an alternative embodiment, the ma- ture polypeptide is amino acids 1 to 315 of SEQ ID NO: 32.

It is known in the art that a host cell may produce a mixture of two of more different mature pol- ypeptides (/^'.e., with a different C-terminal and/or N-terminal amino acid) expressed by the same polynucleotide. It is also known in the art that different host cells process polypeptides different- ly, and thus, one host cell expressing a polynucleotide may produce a different mature polypep- tide (e.g., having a different C-terminal and/or N-terminal amino acid) as compared to another host cell expressing the same polynucleotide.

Mature polypeptide coding sequence: The term “mature polypeptide coding sequence” means a polynucleotide that encodes a mature polypeptide having galactanase activity. Nucleic acid construct: The term "nucleic acid construct" means a nucleic acid molecule, ei- ther single- or double-stranded, which is isolated from a naturally occurring gene or is modified to contain segments of nucleic acids in a manner that would not otherwise exist in nature or which is synthetic, which comprises one or more control sequences.

Nutrient Digestibility: The term“nutrient digestibility” means the fraction of a nutrient that dis appears from the gastro-intestinal tract or a specified segment of the gastro-intestinal tract, e.g. the small intestine. Nutrient digestibility may be measured as the difference between what is administered to the subject and what comes out in the faeces of the subject, or between what is administered to the subject and what remains in the digesta on a specified segment of the gas- tro intestinal tract, e.g. the ileum.

Nutrient digestibility as used herein may be measured by the difference between the intake of a nutrient and the excreted nutrient by means of the total collection of excreta during a period of time; or with the use of an inert marker that is not absorbed by the animal, and allows the re- searcher calculating the amount of nutrient that disappeared in the entire gastro-intestinal tract or a segment of the gastro-intestinal tract. Such an inert marker may be titanium dioxide, chro- mic oxide or acid insoluble ash. Digestibility may be expressed as a percentage of the nutrient in the feed, or as mass units of digestible nutrient per mass units of nutrient in the feed. Nutrient digestibility as used herein encompasses starch digestibility, fat digestibility, protein digestibility, and amino acid digestibility.

Energy digestibility as used herein means the gross energy of the feed consumed minus the gross energy of the faeces or the gross energy of the feed consumed minus the gross energy of the remaining digesta on a specified segment of the gastro-intestinal tract of the animal, e.g. the ileum. Metabolizable energy as used herein refers to apparent metabolizable energy and means the gross energy of the feed consumed minus the gross energy contained in the faeces, urine, and gaseous products of digestion. Energy digestibility and metabolizable energy may be measured as the difference between the intake of gross energy and the gross energy excreted in the faeces or the digesta present in specified segment of the gastro-intestinal tract using the same methods to measure the digestibility of nutrients, with appropriate corrections for nitrogen excretion to calculate metabolizable energy of feed.

Operably linked: The term“operably linked” means a configuration in which a control sequence is placed at an appropriate position relative to the coding sequence of a polynucleotide such that the control sequence directs expression of the coding sequence.

Sequence Identity: The relatedness between two amino acid sequences or between two nu- cleotide sequences is described by the parameter“sequence identity”.

For purposes of the present invention, the degree of sequence identity between two amino acid sequences is determined using the Needleman-Wunsch algorithm (Needleman and Wunsch, 1970, J. Mol. Biol. 48: 443-453) as implemented in the Needle program of the EMBOSS pack- age (EMBOSS: The European Molecular Biology Open Software Suite, Rice et al., 2000, Trends Genet. 16: 276-277), preferably version 3.0.0 or later. Version 6.1.0 was used. The op- tional parameters used are gap open penalty of 10, gap extension penalty of 0.5, and the EBLOSUM62 (EMBOSS version of BLOSUM62) substitution matrix. The output of Needle la- belled“longest identity” (obtained using the -nobrief option) is used as the percent identity and is calculated as follows:

(Identical Residues x 100)/(Length of Alignment - Total Number of Gaps in Alignment)

For purposes of the present invention, the degree of sequence identity between two deoxyribo- nucleotide sequences is determined using the Needleman-Wunsch algorithm (Needleman and Wunsch, 1970, supra) as implemented in the Needle program of the EMBOSS package (EM- BOSS: The European Molecular Biology Open Software Suite, Rice et a!., 2000, supra), prefer- ably version 3.0.0 or later. Version 6.1.0 was used. The optional parameters used are gap open penalty of 10, gap extension penalty of 0.5, and the EDNAFULL (EMBOSS version of NCBI NUC4.4) substitution matrix. The output of Needle labelled“longest identity” (obtained using the -nobrief option) is used as the percent identity and is calculated as follows:

(Identical Deoxyribonucleotides x 100)/(Length of Alignment - Total Number of Gaps in Align ment)

Soy-based material:“soy-based material” means material derived from a Soybean plant such as e.g. dehulled full-fat flakes, full-fat soybean meal, defatted flakes, white flakes, full-fat flakes and soybean meal.

Stringency conditions: The different stringency conditions are defined as follows.

The term“very low stringency conditions” means for probes of at least 100 nucleotides in length, prehybridization and hybridization at 42°C in 5X SSPE, 0.3% SDS, 200 micrograms/ml sheared and denatured salmon sperm DNA, and 25% formamide, following standard Southern blotting procedures for 12 to 24 hours. The carrier material is finally washed three times each for 15 minutes using 2. OX SSC, 0.2% SDS at 60°C.

The term“low stringency conditions” means for probes of at least 100 nucleotides in length, prehybridization and hybridization at 42°C in 5X SSPE, 0.3% SDS, 200 micrograms/ml sheared and denatured salmon sperm DNA, and 25% formamide, following standard Southern blotting procedures for 12 to 24 hours. The carrier material is finally washed three times each for 15 minutes using 1 0X SSC, 0.2% SDS at 60°C.

The term“medium stringency conditions” means for probes of at least 100 nucleotides in length, prehybridization and hybridization at 42°C in 5X SSPE, 0.3% SDS, 200 micrograms/ml sheared and denatured salmon sperm DNA, and 35% formamide, following standard Southern blotting procedures for 12 to 24 hours. The carrier material is finally washed three times each for 15 minutes using 1.OX SSC, 0.2% SDS at 65°C.

The term“medium-high stringency conditions” means for probes of at least 100 nucleotides in length, prehybridization and hybridization at 42°C in 5X SSPE, 0.3% SDS, 200 micrograms/ml sheared and denatured salmon sperm DNA, and 35% formamide, following standard Southern blotting procedures for 12 to 24 hours. The carrier material is finally washed three times each for 15 minutes using 1 0X SSC, 0.2% SDS at 70°C.

The term“high stringency conditions” means for probes of at least 100 nucleotides in length, prehybridization and hybridization at 42°C in 5X SSPE, 0.3% SDS, 200 micrograms/ml sheared and denatured salmon sperm DNA, and 50% formamide, following standard Southern blotting procedures for 12 to 24 hours. The carrier material is finally washed three times each for 15 minutes using 0.5X SSC, 0.2% SDS at 70°C.

The term“very high stringency conditions” means for probes of at least 100 nucleotides in length, prehybridization and hybridization at 42°C in 5X SSPE, 0.3% SDS, 200 micrograms/ml sheared and denatured salmon sperm DNA, and 50% formamide, following standard Southern blotting procedures for 12 to 24 hours. The carrier material is finally washed three times each for 15 minutes using 0.5X SSC, 0.2% SDS at 75°C.

Subsequence: The term“subsequence” means a polynucleotide having one or more (e.g., several) nucleotides absent from the 5' and/or 3' end of a mature polypeptide coding sequence; wherein the subsequence encodes a fragment having galactanase activity.

Substantially pure polypeptide: The term“substantially pure polypeptide” means a prepara- tion that contains at most 10%, at most 8%, at most 6%, at most 5%, at most 4%, at most 3%, at most 2%, at most 1 %, and at most 0.5% by weight of other polypeptide material with which it is natively or recombinantly associated. Preferably, the polypeptide is at least 92% pure, e.g., at least 94% pure, at least 95% pure, at least 96% pure, at least 97% pure, at least 98% pure, at least 99%, at least 99.5% pure, and 100% pure by weight of the total polypeptide material pre- sent in the preparation. The polypeptides of the present invention are preferably in a substan- tially pure form. This can be accomplished, for example, by preparing the polypeptide by well- known recombinant methods or by classical purification methods.

Variant: The term“variant” means a polypeptide having galactanase activity comprising an al- teration, i.e., a substitution, insertion, and/or deletion of one or more (several) amino acid resi- dues at one or more (several) positions. A substitution means a replacement of an amino acid occupying a position with a different amino acid; a deletion means removal of an amino acid oc- cupying a position; and an insertion means adding 1-3 amino acids adjacent to an amino acid occupying a position. In one embodiment, the variants of the present invention has a higher ac- tivity on soybean meal at pH 5 compared to the benchmark Galactanases from H. insolens and B. licheniformis (determined as described in Example 3) such as an activity of at least 0.7, such as at least 0.8, such as at least 09, such as at least 1.0, such as at least 1.1 , such as at least 1.2, such as at least 1.3, such as at least 1.4 or such as at least 1.5 (Abs 405nm; determined as described in Example 3).

Nomenclature

For purposes of the present invention, the nomenclature [Y/F] means that the amino acid at this position may be a tyrosine (Try, Y) or a phenylalanine (Phe, F). Likewise the nomenclature [V/G/A/l] means that the amino acid at this position may be a valine (Val, V), glycine (Gly, G), alanine (Ala, A) or isoleucine (lie, I), and so forth for other combinations as described herein. Unless otherwise limited further, the amino acid X is defined such that it may be any of the 20 natural amino acids.

DETAILED DESCRIPTION OF THE INVENTION

Method of modifying and/or improving the nutritional value of a soy-based material (first aspect)

In a first aspect the invention relates to a method for modifying and/or improving the nutritional value of a soy-based material, the method comprising the steps of:

i) providing a soy-based material and

ii) treating the soy-based material with a polypeptide having galactanase activity selected from the group consisting of:

(a) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least

87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 3;

(b) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least

87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 7;

(c) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least

87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 11 ;

(d) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least

87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 15;

(e) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least

87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 19;

(f) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least

87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 23;

(g) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least

87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 31 ;

(h) a polypeptide encoded by a polynucleotide that hybridizes under high stringency conditions, or very high stringency conditions with

(i) the mature polypeptide coding sequence of SEQ ID NO: 1 or the cDNA sequence thereof,

(ii) the mature polypeptide coding sequence of SEQ ID NO: 5 or the cDNA sequence thereof,

(iii) the mature polypeptide coding sequence of SEQ ID NO: 9 or the cDNA sequence thereof,

(iv) the mature polypeptide coding sequence of SEQ ID NO: 13 or the cDNA sequence thereof,

(v) the mature polypeptide coding sequence of SEQ ID NO: 17 or the cDNA sequence thereof,

(vi) the mature polypeptide coding sequence of SEQ ID NO: 21 or the cDNA sequence thereof,

(vii) the mature polypeptide coding sequence of SEQ ID NO: 29 or the cDNA sequence thereof, (viii) the full-length complementary strand of (i), (ii), (iii), (iv), (v), (vi) or (vii);

(i) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 1 ;

(j) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 5;

(k) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 9;

(l) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 13;

(m) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 17;

(n) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 21 ;

(o) a variant of SEQ ID NO: 3 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions; (p) a variant of SEQ ID NO: 7 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(q) a variant of SEQ ID NO: 11 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(r) a variant of SEQ ID NO: 15 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(s) a variant of SEQ ID NO: 19 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(t) a variant of SEQ ID NO: 23 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(u) a variant of SEQ ID NO: 31 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13 ,14, 15 or 16 positions;

(v) a polypeptide comprising the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (I), (m), (n), (o), (p), (q), (r), (s), (t) or (u) and a N-terminal and/or C- terminal His-tag and/or HQ-tag; and (w) a fragment of the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (I), (m), (n), (o), (p), (q), (r), (s), (t), (u) or (v) having galactanase activity and having at least 90% of the length of the mature polypeptide, wherein the polypeptide optionally has galactanase activity which is at least 60%, such as at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95% or at least 100% of the galactanase activity of the polypeptide of SEQ ID NO: 3.

The soy based material treated with a polypeptide having galactanase activity as described above can be used as an animal feed or an animal feed additive. The soy based material treated with a polypeptide having galactanase activity as described above can in one embodiment be further processed prior to use as an animal feed or an animal feed additive. The further processing can e.g. comprise a drying step. In one embodiment, the method further comprises the step iii) mixing the soy based material from step ii) with one or more vitamins and/or minerals. In one embodiment, the method further comprises the step iii) mixing the soy based material from step ii) with a cereal such as barley, wheat, rye, oat, maize (corn), rice, and/or sorghum.

In one embodiment of the first aspect the soy-based material is selected from the group consisting of dehulled full-fat flakes, full-fat soybean meal, defatted flakes, white flakes, full-fat flakes and soybean meal. In a preferred embodiment the soy-based material is in an aqueous solution. The soy-based material has in one embodiment a dry-matter content of from 40-60% (w/w) such as from 40%-45% (w/w), for example from 45%-50 (w/w) such as from 50%-55% (w/w), for example from 55%-60 (w/w), or any combination of these intervals. In another preferred embodiment the soy-based material has a dry-matter content of from 20-40% (w/w) such as from 20-25% (w/w), for example from 25-30% (w/w), such as from 30-35% (w/w), for example from 35-40% (w/w) or any combination of these intervals.

In one embodiment of the first aspect step ii) comprises galactanase treatment of the soy-based material for 24 hours to 5 days such as for example from 24 hours to 2 days, such as from 2 days to 3 days, for example from 3 day to 4 days, such as from 4 days to 5 days, or any combination of these intervals. In another preferred embodiment step ii) comprises galactanase treatment of the soy-based material for 6 hours to 24 hours, such as for example for 6 hours to 8 hours, such as for 8 hours to 10 hours, for example for 10 hours to 12 hours, such as for 12 hours to 14 hours, for example for 14 hours to 16 hours, such as for 16 hours to 18 hours, for example for 18 hours to 20 hours, such as for 20 hours to 22 hours, for example for 22 hours to 24 hours, or any combination of these intervals. In a further embodiment step ii) comprises galactanase treatment of the soy-based material for 30 minutes to 6 hours, such as from 30 minutes to 1 hour, for example from 1 hour to 2 hours, such as from 2 hours to 3 hours, for example from 3 hours to 4 hours, such as from 4 hours to 5 hours, for example from 5 hours to 6 hours, or any combination of these intervals. Step ii) of the first aspect comprises in one embodiment use of from 0.001 mg galactanase /dry- matter to 6 mg galactanase /dry-matter such as from 0.001 mg galactanase /dry-matter to 0.005 mg galactanase /dry-matter, for example from 0.005 mg galactanase /dry-matter to 0.01 mg galactanase /dry-matter, such as from 0.01 mg galactanase /dry-matter to 0.02 mg galactanase /dry-matter, for example from 0.02 mg galactanase /dry-matter to 0.05 mg galactanase /dry- matter, such as from 0.05 mg galactanase /dry-matter to 0.1 mg galactanase /dry-matter, for example from 0.1 mg galactanase /dry-matter to 0.5 mg galactanase /dry-matter, such as from 0.5 mg galactanase /dry-matter to 1 mg galactanase /dry-matter, for example from 1 mg galactanase /dry-matter to 2 mg galactanase /dry-matter, such as from 2 mg galactanase /dry- matter to 3 mg galactanase /dry-matter, for example from 3 mg galactanase /dry-matter to 4 mg galactanase /dry-matter, such as from 4 mg galactanase /dry-matter to 5 mg galactanase /dry- matter, for example from 5 mg galactanase /dry-matter to 6 mg galactanase /dry-matter or any combination of these intervals.

In a preferred embodiment step ii) of the first aspect is performed in vitro. In vitro means outside the body of an animal such as outside the gut of an animal. Accordingly, step ii) can be performed in any suitable tank, container, flask, box or the like.

In a specific embodiment step ii) is performed before one or more alcohol leaching steps. In another specific embodiment an alcohol leaching step is performed after step ii).

In a preferred embodiment the galactanase treated soy-based material obtained in the first aspect has an increased protein content (compared to soy-based material that has not been treated with galactanase). Preferably the protein content is increased by at least 1% such as at least 2%, such as at least 3%, such as at least 4%, such as at least 5%, such as at least 6%, such as at least 7%, such as at least 8%, such as at least 9%, or such as at least 10% (compared to soy-based material that has not been treated with galactanase).

In another preferred embodiment the galactanase treated soy-based material obtained in the first aspect has an increased NDF (Neutral Detergent Fiber) digestibility (compared to soy- based material that has not been treated with galactanase). Preferably the NDF digestibility is increased by at least 1 % such as at least 2%, such as at least 3%, such as at least 4%, such as at least 5%, such as at least 6%, such as at least 7%, such as at least 8%, such as at least 9%, or such as at least 10% (compared to soy-based material that has not been treated with galactanase).

Step ii) of the first aspect comprises in one embodiment galactanase treatment of the soy-based material at pH between pH 3 and 6, such as between pH 3 and 4, for example between pH 4 and 5, such as pH 5 and 6, or any combination of these intervals.

Step ii) of the first aspect comprises in one embodiment galactanase treatment of the soy-based material at a temperature between 40°C and 80°C such as between 40°C and 50°C, for example between 50°C and 60°C, such as between 60°C and 70°C, for example between 70°C and 80°C, or any combination of these intervals. Step ii) of the first aspect comprises in one embodiment galactanase treatment of the soy-based material at a temperature between 15°C and 40°C such as between 15°C and 20°C, for example between 20°C and 25°C, such as between 25°C and 30°C, for example between 30°C and 35°C, such as between 35°C and 40°C, or any combination of these intervals.

Step ii) of the first aspect comprises in one embodiment comprises galactanase treatment and fermentation in a single step. Step ii) is in another embodiment followed by one or more fermentation steps.

The method according to the first aspect results e.g. in production of soybean meal, fermented soybean meal or soy protein concentrate.

In a preferred embodiment the polypeptide defined in the first aspect comprises or consists of amino acids 1 to 316 of SEQ ID NO: 2, amino acids 1 to 316 of SEQ ID NO: 3, amino acids 1 to 324 of SEQ ID NO: 4, amino acids 1 to 318 of SEQ ID NO: 6, amino acids 1 to 318 of SEQ ID NO: 7, amino acids 1 to 326 of SEQ ID NO: 8, amino acids 1 to 516 of SEQ ID NO: 10, amino acids 1 to 516 of SEQ ID NO: 1 1 , amino acids 1 to 524 of SEQ ID NO: 12, amino acids 1 to 317 of SEQ ID NO: 14, amino acids 1 to 317 of SEQ ID NO: 15, amino acids 1 to 325 of SEQ ID NO: 16, amino acids 1 to 316 of SEQ ID NO: 18, amino acids 1 to 316 of SEQ ID NO: 19, amino acids 1 to 324 of SEQ ID NO: 20, amino acids 1 to 316 of SEQ ID NO: 22, amino acids 1 to 316 of SEQ ID NO: 23, amino acids 1 to 324 of SEQ ID NO: 24, amino acids 1 to 307 of SEQ ID NO: 30, amino acids 1 to 307 of SEQ ID NO: 31 or amino acids 1 to 315 of SEQ ID NO: 24.

The invention also relates to an animal feed or animal feed additive comprising the galactanase treated soy-based material obtained in the method according to the first aspect described above. The invention also relates to use the galactanase treated soy-based material obtained in the method according to the first aspect as an animal feed or animal feed additive.

Polypeptides Having Galactanase Activity (Second to Seventh aspect)

In a second aspect, the invention relates to polypeptides having galactanase activity and having a sequence identity to the mature polypeptide of SEQ ID NO: 2 of at least 82%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, which have galactanase activity. In one embodiment, the polypeptides differ by up to 50 amino acids, e.g., between 1 and 50 amino acids, such as 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1- 15, 1-10 or 1-5 amino acids, or 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 amino acids from the mature polypeptide of SEQ ID NO: 2.

In one embodiment, the polypeptide preferably comprises or consists of the amino acid se- quence of SEQ ID NO: 2 or an allelic variant thereof; comprises the amino acid sequence of SEQ ID NO: 2 and a N-terminal and/or C-terminal His-tag and/or HQ-tag; or is a fragment thereof having galactanase activity and having at least 90% such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% of the length of the mature polypeptide. In another embodiment, the polypeptide comprises or consists of the mature polypeptide of SEQ ID NO: 2. In another embodiment, the polypeptide comprises or consists of amino acids 1 to 316 of SEQ ID NO: 2. In an embodiment, the polypep- tide has been isolated.

In a continuation of the second aspect, the invention further relates to polypeptides having ga- lactanase activity having a sequence identity to SEQ ID NO: 3 of at least 82%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 3 of at least 85%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 3 of at least 86%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 3 of at least 87%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 3 of at least 88%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 3 of at least 89%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 3 of at least 90%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 3 of at least 91 %. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 3 of at least 92%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 3 of at least 93%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 3 of at least 94%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 3 of at least 95%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 3 of at least 96%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 3 of at least 97%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 3 of at least 98%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 3 of at least 99%.

In one embodiment, the polypeptides differ by up to 50 amino acids, e.g., between 1 and 50 amino acids, such as 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10 or 1-5 amino acids, or 1 , 2,

3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29,

30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 amino acids from SEQ ID NO: 3. In one embodiment, the polypeptides differ by up to 50 amino acids, e.g., between 1 and 50 amino acids, such as 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10 or 1-5 amino acids, or 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or

50 amino acids from SEQ ID NO: 4.

In one embodiment, the polypeptide preferably comprises or consists of the amino acid se- quence of SEQ ID NO: 3 or an allelic variant thereof; comprises the amino acid sequence of SEQ ID NO: 3 and a N-terminal and/or C-terminal His-tag and/or HQ-tag such as SEQ ID NO: 4; or is a fragment thereof having galactanase activity and having at least 90% such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% of the length of the mature polypeptide. In another embodiment, the poly- peptide comprises or consists of amino acids 1 to 316 of SEQ ID NO: 3. In another embodi- ment, the polypeptide comprises or consists of amino acids 1 to 324 of SEQ ID NO: 4. In an embodiment, the polypeptide has been isolated.

In a continuation of the second aspect, the invention relates to a polypeptide having galac- tanase activity encoded by a polynucleotide that hybridizes under high or very high stringency conditions with (i) the mature polypeptide coding sequence of SEQ ID NO: 1 , (ii) the cDNA se- quence thereof, or (iii) the full-length complement of (i) or (ii) (Sambrook ef a/., 1989, Molecular Cloning, A Laboratory Manual, 2d edition, Cold Spring Harbor, New York). In an embodiment, the polypeptide has been isolated.

In a continuation of the second aspect, the invention relates to a polypeptide having galac- tanase activity encoded by a polynucleotide having a sequence identity to the mature polypep- tide coding sequence of SEQ ID NO: 1 or the cDNA sequence thereof of at least 82%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%. In a further embodiment, the polypeptide has been isolated.

In a continuation of the second aspect, the invention relates to variants of SEQ ID NO: 3 having galactanase activity comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof at one or more (e.g., several) positions. In an embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in SEQ ID NO: 3 is not more than 50, e.g., 1 ,

2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29,

30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50. In another embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in SEQ ID NO: 3 is between 1 and 48, such as 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10 or 1-5 positions. In an embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid in- sertions or any combination thereof in SEQ ID NO: 3 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10. In another embodiment, the number of substitutions, deletions, and/or insertions in SEQ ID NO: 3 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further embodiment, the number of substitutions in SEQ ID NO: 3 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further embodiment, the number of conservative substitutions in SEQ ID NO: 3 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10.

The amino acid changes may be of a minor nature, that is conservative amino acid substitutions or insertions that do not significantly affect the folding and/or activity of the protein; small dele- tions, typically of 1-30 amino acids; small amino- or carboxyl-terminal extensions, such as an amino-terminal methionine residue; a small linker peptide of up to 20-25 residues; or a small extension that facilitates purification by changing net charge or another function, such as a poly- histidine tract, an antigenic epitope or a binding domain.

Examples of conservative substitutions are within the groups of basic amino acids (arginine, ly- sine and histidine), acidic amino acids (glutamic acid and aspartic acid), polar amino acids (glu tamine and asparagine), hydrophobic amino acids (leucine, isoleucine and valine), aromatic amino acids (phenylalanine, tryptophan and tyrosine), and small amino acids (glycine, alanine, serine, threonine and methionine). Amino acid substitutions that do not generally alter specific activity are known in the art and are described, for example, by H. Neurath and R.L. Hill, 1979, In, The Proteins, Academic Press, New York. Common substitutions are Ala/Ser, Val/lle, Asp/Glu, Thr/Ser, Ala/Gly, Ala/Thr, Ser/Asn, Ala/Val, Ser/Gly, Tyr/Phe, Ala/Pro, Lys/Arg, Asp/Asn, Leu/lle, Leu/Val, Ala/Glu, and Asp/Gly. Other examples of conservative substitutions are G to A; A to G, S; V to I, L, A, T, S; I to V, L, M; L to I, M, V; M to L, I, V; P to A, S, N; F to Y, W, H; Y to F, W, H; W to Y, F, H; R to K, E, D; K to R, E, D; H to Q, N, S; D to N, E, K, R, Q; E to Q, D, K, R, N; S to T, A; T to S, V, A; C to S, T, A; N to D, Q, H, S; Q to E, N, H, K, R.

Alternatively, the amino acid changes are of such a nature that the physico-chemical properties of the polypeptides are altered. For example, amino acid changes may improve the thermal sta- bility of the polypeptide, alter the substrate specificity, change the pH optimum, and the like.

Essential amino acids in a polypeptide can be identified according to procedures known in the art, such as site-directed mutagenesis or alanine-scanning mutagenesis (Cunningham and Wells, 1989, Science 244: 1081-1085). In the latter technique, single alanine mutations are in- troduced at every residue in the molecule, and the resultant mutant molecules are tested for ga- lactanase activity to identify amino acid residues that are critical to the activity of the molecule. See also, Hilton et el., 1996, J. Biol. Chem. 271 : 4699-4708. The active site of the enzyme or other biological interaction can also be determined by physical analysis of structure, as deter- mined by such techniques as nuclear magnetic resonance, crystallography, electron diffraction, or photoaffinity labelling, in conjunction with mutation of putative contact site amino acids. See, for example, de Vos et el., 1992, Science 255: 306-312; Smith et el., 1992, J. Mol. Biol. 224: 899-904; Wlodaver et el., 1992, FEBS Lett. 309: 59-64. The identity of essential amino acids can also be inferred from an alignment with a related polypeptide.

Single or multiple amino acid substitutions, deletions, and/or insertions can be made and tested using known methods of mutagenesis, recombination, and/or shuffling, followed by a relevant screening procedure, such as those disclosed by Reidhaar-Olson and Sauer, 1988, Science 241 : 53-57; Bowie and Sauer, 1989, Proc. Natl. Acad. Sci. USA 86: 2152-2156; WO 95/17413; or WO 95/22625. Other methods that can be used include error-prone PCR, phage display (e.g., Lowman et al., 1991 , Biochemistry 30: 10832-10837; U.S. Patent No. 5,223,409; WO 92/06204), and region-directed mutagenesis (Derbyshire et al., 1986, Gene 46: 145; Ner et a!., 1988, DNA 7: 127).

Mutagenesis/shuffling methods can be combined with high-throughput, automated screening methods to detect activity of cloned, mutagenized polypeptides expressed by host cells (Ness et al., 1999, Nature Biotechnology 17: 893-896). Mutagenized DNA molecules that encode active polypeptides can be recovered from the host cells and rapidly sequenced using standard meth- ods in the art. These methods allow the rapid determination of the importance of individual ami- no acid residues in a polypeptide.

The polypeptide may be a hybrid polypeptide in which a region of one polypeptide is fused at the N-terminus or the C-terminus of a region of another polypeptide.

The polypeptide may be a fusion polypeptide or cleavable fusion polypeptide in which another polypeptide is fused at the N-terminus or the C-terminus of the polypeptide of the present inven- tion. A fusion polypeptide is produced by fusing a polynucleotide encoding another polypeptide to a polynucleotide of the present invention. Techniques for producing fusion polypeptides are known in the art, and include ligating the coding sequences encoding the polypeptides so that they are in frame and that expression of the fusion polypeptide is under control of the same promoter(s) and terminator. Fusion polypeptides may also be constructed using intein technolo- gy in which fusion polypeptides are created post-translationally (Cooper et al., 1993, EMBO J. 12: 2575-2583; Dawson et a!., 1994, Science 266: 776-779).

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

Carbohydrate molecules are often attached to a polypeptide from a fungal source during post- translational modification. In order to aid mass spectrometry analysis, the polypeptide can be incubated with an endoglycosidase to deglycosylate each /V-linked position. For every deglyco- sylated /V-linked site, one N- acetyl hexosamine remains on the protein backbone.

In an embodiment, the polypeptide of the second aspect comprises the motif GV[T/M]PD[W/M]VQ[I/V]GNE (SEQ ID NO: 26) and/or the motif WADP[A/G]xQxKPxAW (SEQ ID NO: 27). In a third aspect, the invention relates to polypeptides having galactanase activity and having a sequence identity to the mature polypeptide of SEQ ID NO: 6 of at least 83%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, which have galactanase activity. In one embodiment, the polypeptides differ by up to 50 amino acids, e.g., between 1 and 50 amino acids, such as 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1- 10 or 1-5 amino acids, or 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 ,

22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46,

47, 48, 49 or 50 amino acids from the mature polypeptide of SEQ ID NO: 6.

In one embodiment, the polypeptide preferably comprises or consists of the amino acid se- quence of SEQ ID NO: 6 or an allelic variant thereof; comprises the amino acid sequence of SEQ ID NO: 6 and a N-terminal and/or C-terminal His-tag and/or HQ-tag; or is a fragment thereof having galactanase activity and having at least 90% such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% of the length of the mature polypeptide. In another embodiment, the polypeptide comprises or consists of the mature polypeptide of SEQ ID NO: 6. In another embodiment, the polypeptide comprises or consists of amino acids 1 to 318 of SEQ ID NO: 6. In an embodiment, the polypep- tide has been isolated.

In a continuation of the third aspect, the invention further relates to polypeptides having galac- tanase activity having a sequence identity to SEQ ID NO : 7 of at least 83%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 7 of at least 85%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 7 of at least 86%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 7 of at least 87%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 7 of at least 88%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 7 of at least 89%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 7 of at least 90%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 7 of at least 91%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 7 of at least 92%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 7 of at least 93%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 7 of at least 94%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 7 of at least 95%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 7 of at least 96%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 7 of at least 97%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 7 of at least 98%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 7 of at least 99%.

In one embodiment, the polypeptides differ by up to 50 amino acids, e.g., between 1 and 50 amino acids, such as 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10 or 1-5 amino acids, or 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 amino acids from SEQ ID NO: 7. In one embodiment, the polypeptides differ by up to 50 amino acids, e.g., between 1 and 50 amino acids, such as 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10 or 1-5 amino acids, or 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 amino acids from SEQ ID NO: 8.

In one embodiment, the polypeptide preferably comprises or consists of the amino acid se- quence of SEQ ID NO: 7 or an allelic variant thereof; comprises the amino acid sequence of SEQ ID NO: 7 and a N-terminal and/or C-terminal His-tag and/or HQ-tag such as SEQ ID NO: 8; or is a fragment thereof having galactanase activity and having at least 90% such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% of the length of the mature polypeptide. In another embodiment, the poly- peptide comprises or consists of amino acids 1 to 318 of SEQ ID NO: 7. In another embodi- ment, the polypeptide comprises or consists of amino acids 1 to 326 of SEQ ID NO: 8. In an embodiment, the polypeptide has been isolated.

In a continuation of the third aspect, the invention relates to a polypeptide having galactanase activity encoded by a polynucleotide that hybridizes under high or very high stringency condi- tions with (i) the mature polypeptide coding sequence of SEQ ID NO: 5, (ii) the cDNA sequence thereof, or (iii) the full-length complement of (i) or (ii) (Sambrook et al., supra). In an embodi- ment, the polypeptide has been isolated.

In a continuation of the third aspect, the invention relates to a polypeptide having galactanase activity encoded by a polynucleotide having a sequence identity to the mature polypeptide cod- ing sequence of SEQ ID NO: 5 or the cDNA sequence thereof of at least 83%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%. In a further embodiment, the polypeptide has been isolated.

In a continuation of the third aspect, the invention relates to variants of SEQ ID NO: 7 having galactanase activity comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof at one or more (e.g., several) positions. In an embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in SEQ ID NO: 7 is not more than 50, e.g., 1 ,

30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50. In another embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in SEQ ID NO: 7 is between 1 and 48, such as 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10 or 1-5 positions. In an embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid in- sertions or any combination thereof in SEQ ID NO: 7 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10. In another embodiment, the number of substitutions, deletions, and/or insertions in SEQ ID NO: 7 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further embodiment, the number of substitutions in SEQ ID NO: 7 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further embodiment, the number of conservative substitutions in SEQ ID NO: 7 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10.

Examples of amino acid changes and conservative substitutions are described in the second aspect of the invention.

In an embodiment, the polypeptide of the third aspect comprises the motif GV[T/M]PD[W/M]VQ[IA/]GNE (SEQ ID NO: 26) and/or the motif WADP[A/G]xQxKPxAW (SEQ ID NO: 27).

In a fourth aspect, the invention relates to polypeptides having galactanase activity and having a sequence identity to the mature polypeptide of SEQ ID NO: 10 of at least 86%, e.g., at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, which have galac- tanase activity. In one embodiment, the polypeptides differ by up to 50 amino acids, e.g., be- tween 1 and 50 amino acids, such as 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10 or 1-5 ami- no acids, or 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25,

26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 amino acids from the mature polypeptide of SEQ ID NO: 10.

In one embodiment, the polypeptide preferably comprises or consists of the amino acid se- quence of SEQ ID NO: 10 or an allelic variant thereof; comprises the amino acid sequence of SEQ ID NO: 10 and a N-terminal and/or C-terminal His-tag and/or HQ-tag; or is a fragment thereof having galactanase activity and having at least 90% such as at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% of the length of the mature polypeptide. In another embodiment, the polypeptide comprises or consists of the mature polypeptide of SEQ ID NO: 10. In another embodiment, the polypep- tide comprises or consists of amino acids 1 to 516 of SEQ ID NO: 10. In an embodiment, the polypeptide has been isolated.

In a continuation of the fourth aspect, the invention further relates to polypeptides having galac- tanase activity having a sequence identity to SEQ ID NO: 1 1 of at least 86%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 11 of at least 87%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 11 of at least 88%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 11 of at least 89%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 11 of at least 90%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 11 of at least 91 %. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 11 of at least 92%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 11 of at least 93%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 11 of at least 94%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 11 of at least 95%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 11 of at least 96%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 11 of at least 97%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 11 of at least 98%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 11 of at least 99%.

30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 amino acids from SEQ ID NO: 11. In one embodiment, the polypeptides differ by up to 50 amino acids, e.g., between 1 and 50 amino acids, such as 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10 or 1-5 amino acids, or 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or

50 amino acids from SEQ ID NO: 12.

In one embodiment, the polypeptide preferably comprises or consists of the amino acid se- quence of SEQ ID NO: 11 or an allelic variant thereof; comprises the amino acid sequence of SEQ ID NO: 1 1 and a N-terminal and/or C-terminal His-tag and/or HQ-tag such as SEQ ID NO: 12; or is a fragment thereof having galactanase activity and having at least 90% such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% of the length of the mature polypeptide. In another embodiment, the poly- peptide comprises or consists of amino acids 1 to 516 of SEQ ID NO: 1 1. In another embodi- ment, the polypeptide comprises or consists of amino acids 1 to 524 of SEQ ID NO: 12. In an embodiment, the polypeptide has been isolated.

In a continuation of the fourth aspect, the invention relates to a polypeptide having galactanase activity encoded by a polynucleotide that hybridizes under high stringency conditions or very high stringency conditions with (i) the mature polypeptide coding sequence of SEQ ID NO: 9, (ii) the cDNA sequence thereof, or (iii) the full-length complement of (i) or (ii) (Sambrook et ah, su pra). In an embodiment, the polypeptide has been isolated.

In a continuation of the fourth aspect, the invention relates to a polypeptide having galactanase activity encoded by a polynucleotide having a sequence identity to the mature polypeptide cod- ing sequence of SEQ ID NO: 9 or the cDNA sequence thereof of at least 86%, e.g., at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%. In a further embodiment, the polypeptide has been isolated.

In a continuation of the fourth aspect, the invention relates to variants of SEQ ID NO: 1 1 having galactanase activity comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof at one or more (e.g., several) positions. In an embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in SEQ ID NO: 11 is not more than 50, e.g., 1 ,

30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50. In another embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in SEQ ID NO: 11 is between 1 and 48, such as 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10 or 1-5 positions. In an embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid in- sertions or any combination thereof in SEQ ID NO: 1 1 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10. In another embodiment, the number of substitutions, deletions, and/or insertions in SEQ ID NO: 1 1 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further embodiment, the number of substitutions in SEQ ID NO: 11 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further embodiment, the number of conservative substitutions in SEQ ID NO: 1 1 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10.

In an embodiment, the polypeptide of the fourth aspect comprises the motif GV[T/M]PD[W/M]VQ[I/V]GNE (SEQ ID NO: 26) and/or the motif WADP[A/G]xQxKPxAW (SEQ ID NO: 27).

In a fifth aspect, the invention relates to polypeptides having galactanase activity and having a sequence identity to the mature polypeptide of SEQ ID NO: 14 of at least 99.3%, e.g., at least 99.6% which have galactanase activity. In one embodiment, the polypeptides differ by up to 2 amino acids, e.g., 1 or 2 amino acids from the mature polypeptide of SEQ ID NO: 14.

In one embodiment, the polypeptide preferably comprises or consists of the amino acid se- quence of SEQ ID NO: 14 or an allelic variant thereof; comprises the amino acid sequence of SEQ ID NO: 14 and a N-terminal and/or C-terminal His-tag and/or HQ-tag; or is a fragment thereof having galactanase activity and having at least 90% such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% of the length of the mature polypeptide. In another embodiment, the polypeptide comprises or consists of the mature polypeptide of SEQ ID NO: 14. In another embodiment, the polypep- tide comprises or consists of amino acids 1 to 317 of SEQ ID NO: 14. In an embodiment, the polypeptide has been isolated.

In a continuation of the fifth aspect, the invention further relates to polypeptides having galac- tanase activity having a sequence identity to SEQ ID NO: 15 of at least 99.3%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 15 of at least 99.6%.

In one embodiment, the polypeptides differ by up to 2 amino acids, e.g., 1 or 2 amino acids from SEQ ID NO: 15. In one embodiment, the polypeptides differ by up to 2 amino acids, e.g., 1 or 2 amino acids from SEQ ID NO: 16.

In one embodiment, the polypeptide preferably comprises or consists of the amino acid se- quence of SEQ ID NO: 15 or an allelic variant thereof; comprises the amino acid sequence of SEQ ID NO: 15 and a N-terminal and/or C-terminal His-tag and/or HQ-tag such as SEQ ID NO: 16; or is a fragment thereof having galactanase activity and having at least 90% such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% of the length of the mature polypeptide. In another embodiment, the poly- peptide comprises or consists of amino acids 1 to 317 of SEQ ID NO: 15. In another embodi- ment, the polypeptide comprises or consists of amino acids 1 to 325 of SEQ ID NO: 16. In an embodiment, the polypeptide has been isolated.

In a continuation of the fifth aspect, the invention relates to a polypeptide having galactanase activity encoded by a polynucleotide having a sequence identity to the mature polypeptide cod- ing sequence of SEQ ID NO: 13 or the cDNA sequence thereof of at least 99.3%, e.g., at least 99.6%, or 100%. In a further embodiment, the polypeptide has been isolated.

In a continuation of the fifth aspect, the invention relates to variants of SEQ ID NO: 15 having galactanase activity comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof at one or more (e.g., several) positions. In an embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in SEQ ID NO: 15 is not more than 2 amino acids, e.g., 1 or 2. In another embodiment, the number of substitutions, deletions, and/or inser- tions in SEQ ID NO: 15 is not more than 2 amino acids, e.g., 1 or 2. In a further embodiment, the number of substitutions in SEQ ID NO: 15 is not more than 2 amino acids, e.g., 1 or 2. In a further embodiment, the number of conservative substitutions in SEQ ID NO: 15 is not more than 2 amino acids, e.g., 1 or 2. Examples of amino acid changes and conservative substitutions are described in the second aspect of the invention.

In an embodiment, the polypeptide of the fifth aspect comprises the motif GV[T/M]PD[W/M]VQ[IA/]GNE (SEQ ID NO: 26) and/or the motif WADP[A/G]xQxKPxAW (SEQ ID NO: 27).

In a sixth aspect, the invention relates to polypeptides having galactanase activity and having a sequence identity to the mature polypeptide of SEQ ID NO: 18 of at least 99.3%, e.g., at least 99.6% which have galactanase activity. In one embodiment, the polypeptides differ by up to 2 amino acids, e.g., 1 or 2 amino acids from the mature polypeptide of SEQ ID NO: 18.

In one embodiment, the polypeptide preferably comprises or consists of the amino acid se- quence of SEQ ID NO: 18 or an allelic variant thereof; comprises the amino acid sequence of SEQ ID NO: 18 and a N-terminal and/or C-terminal His-tag and/or HQ-tag; or is a fragment thereof having galactanase activity and having at least 90% such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% of the length of the mature polypeptide. In another embodiment, the polypeptide comprises or consists of the mature polypeptide of SEQ ID NO: 18. In another embodiment, the polypep- tide comprises or consists of amino acids 1 to 316 of SEQ ID NO: 18. In an embodiment, the polypeptide has been isolated.

In a continuation of the sixth aspect, the invention further relates to polypeptides having galac- tanase activity having a sequence identity to SEQ ID NO: 19 of at least 99.3%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 19 of at least 99.6%.

In one embodiment, the polypeptides differ by up to 2 amino acids, e.g., 1 or 2 amino acids from SEQ ID NO: 19. In one embodiment, the polypeptides differ by up to 2 amino acids, e.g., 1 or 2 amino acids from SEQ ID NO: 20.

In one embodiment, the polypeptide preferably comprises or consists of the amino acid se- quence of SEQ ID NO: 19 or an allelic variant thereof; comprises the amino acid sequence of SEQ ID NO: 19 and a N-terminal and/or C-terminal His-tag and/or HQ-tag such as SEQ ID NO: 20; or is a fragment thereof having galactanase activity and having at least 90% such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% of the length of the mature polypeptide. In another embodiment, the poly- peptide comprises or consists of amino acids 1 to 316 of SEQ ID NO: 19. In another embodi- ment, the polypeptide comprises or consists of amino acids 1 to 324 of SEQ ID NO: 20. In an embodiment, the polypeptide has been isolated.

In a continuation of the sixth aspect, the invention relates to a polypeptide having galactanase activity encoded by a polynucleotide having a sequence identity to the mature polypeptide cod- ing sequence of SEQ ID NO: 17 or the cDNA sequence thereof of at least 99.3%, e.g., at least 99.6%, or 100%. In a further embodiment, the polypeptide has been isolated.

In a continuation of the sixth aspect, the invention relates to variants of SEQ ID NO: 19 having galactanase activity comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof at one or more (e.g., several) positions. In an embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in SEQ ID NO: 19 is not more than 2 amino acids, e.g., 1 or 2. In another embodiment, the number of substitutions, deletions, and/or inser- tions in SEQ ID NO: 19 is not more than 2 amino acids, e.g., 1 or 2. In a further embodiment, the number of substitutions in SEQ ID NO: 19 is not more than 2 amino acids, e.g., 1 or 2. In a further embodiment, the number of conservative substitutions in SEQ ID NO: 19 is not more than 2 amino acids, e.g., 1 or 2.

In an embodiment, the polypeptide of the sixth aspect comprises the motif GV[T/M]PD[W/M]VQ[I/V]GNE (SEQ ID NO: 26) and/or the motif WADP[A/G]xQxKPxAW (SEQ ID NO: 27).

In a seventh aspect, the invention relates to polypeptides having galactanase activity and hav- ing a sequence identity to the mature polypeptide of SEQ ID NO: 22 of at least 83%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, which have galactanase activity. In one embodiment, the polypeptides differ by up to 50 amino acids, e.g., between 1 and 50 amino acids, such as 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10 or 1-5 amino acids, or 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19,

20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44,

45, 46, 47, 48, 49 or 50 amino acids from the mature polypeptide of SEQ ID NO: 22.

In one embodiment, the polypeptide preferably comprises or consists of the amino acid se- quence of SEQ ID NO: 22 or an allelic variant thereof; comprises the amino acid sequence of SEQ ID NO: 22 and a N-terminal and/or C-terminal His-tag and/or HQ-tag; or is a fragment thereof having galactanase activity and having at least 90% such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% of the length of the mature polypeptide. In another embodiment, the polypeptide comprises or consists of the mature polypeptide of SEQ ID NO: 22. In another embodiment, the polypep- tide comprises or consists of amino acids 1 to 316 of SEQ ID NO: 22. In an embodiment, the polypeptide has been isolated.

In a continuation of the seventh aspect, the invention further relates to polypeptides having ga- lactanase activity having a sequence identity to SEQ ID NO: 23 of at least 83%. In an embodi- ment, the polypeptide has a sequence identity to SEQ ID NO: 23 of at least 85%. In an embod- iment, the polypeptide has a sequence identity to SEQ ID NO: 23 of at least 86%. In an embod- iment, the polypeptide has a sequence identity to SEQ ID NO: 23 of at least 87%. In an embod- iment, the polypeptide has a sequence identity to SEQ ID NO: 23 of at least 88%. In an embod- iment, the polypeptide has a sequence identity to SEQ ID NO: 23 of at least 89%. In an embod- iment, the polypeptide has a sequence identity to SEQ ID NO: 23 of at least 90%. In an embod- iment, the polypeptide has a sequence identity to SEQ ID NO: 23 of at least 91 %. In an embod- iment, the polypeptide has a sequence identity to SEQ ID NO: 23 of at least 92%. In an embod- iment, the polypeptide has a sequence identity to SEQ ID NO: 23 of at least 93%. In an embod- iment, the polypeptide has a sequence identity to SEQ ID NO: 23 of at least 94%. In an embod- iment, the polypeptide has a sequence identity to SEQ ID NO: 23 of at least 95%. In an embod- iment, the polypeptide has a sequence identity to SEQ ID NO: 23 of at least 96%. In an embod- iment, the polypeptide has a sequence identity to SEQ ID NO: 23 of at least 97%. In an embod- iment, the polypeptide has a sequence identity to SEQ ID NO: 23 of at least 98%. In an embod- iment, the polypeptide has a sequence identity to SEQ ID NO: 23 of at least 99%.

30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 amino acids from SEQ ID NO: 23. In one embodiment, the polypeptides differ by up to 50 amino acids, e.g., between 1 and 50 amino acids, such as 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10 or 1-5 amino acids, or 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 amino acids from SEQ ID NO: 24.

In one embodiment, the polypeptide preferably comprises or consists of the amino acid se- quence of SEQ ID NO: 23 or an allelic variant thereof; comprises the amino acid sequence of SEQ ID NO: 23 and a N-terminal and/or C-terminal His-tag and/or HQ-tag such as SEQ ID NO: 24; or is a fragment thereof having galactanase activity and having at least 90% such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% of the length of the mature polypeptide. In another embodiment, the poly- peptide comprises or consists of amino acids 1 to 316 of SEQ ID NO: 23. In another embodi- ment, the polypeptide comprises or consists of amino acids 1 to 324 of SEQ ID NO: 24. In an embodiment, the polypeptide has been isolated. In a continuation of the seventh aspect, the invention relates to a polypeptide having galac- tanase activity encoded by a polynucleotide that hybridizes under high stringency conditions or very high stringency conditions with (i) the mature polypeptide coding sequence of SEQ ID NO: 21 , (ii) the cDNA sequence thereof, or (iii) the full-length complement of (i) or (ii) (Sambrook et a!., supra). In an embodiment, the polypeptide has been isolated.

In a continuation of the seventh aspect, the invention relates to a polypeptide having galac- tanase activity encoded by a polynucleotide having a sequence identity to the mature polypep- tide coding sequence of SEQ ID NO: 21 or the cDNA sequence thereof of at least 83%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%. In a further embodiment, the polypeptide has been isolated.

In a continuation of the seventh aspect, the invention relates to variants of SEQ ID NO: 23 hav- ing galactanase activity comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof at one or more (e.g., several) positions. In an embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in SEQ ID NO: 23 is not more than 50, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27,

28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50. In an- other embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any com- bination thereof in SEQ ID NO: 23 is between 1 and 48, such as 1-45, 1-40, 1-35, 1-30, 1-25, 1- 20, 1-15, 1-10 or 1-5 positions. In an embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in SEQ ID NO: 23 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10. In another embodiment, the number of substitutions, deletions, and/or insertions in SEQ ID NO: 23 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further embodiment, the number of substitutions in SEQ ID NO: 23 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further embodiment, the number of conservative substitutions in SEQ ID NO: 23 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10.

In an embodiment, the polypeptide of the seventh aspect comprises the motif GV[T/M]PD[W/M]VQ[I/V]GNE (SEQ ID NO: 26) and/or the motif WADP[A/G]xQxKPxAW (SEQ ID NO: 27). In an eighth aspect, the invention relates to polypeptides having galactanase activity and having a sequence identity to the mature polypeptide of SEQ ID NO: 30 of at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or at least 99.5%, which have galactanase activity. In one embodiment, the polypeptides differ by up to 16 amino acids, e.g., between 1 and 16 amino acids, such as 1-15, 1-14, 1-13, 1-12, 1-11 , 1-10, 1-9, 1-8, 1-7, 1-6 or 1-5 amino acids, or 1 , 2,

3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15 or 16 amino acids from the mature polypeptide of SEQ ID NO: 30.

In one embodiment, the polypeptide preferably comprises or consists of the amino acid se- quence of SEQ ID NO: 30 or an allelic variant thereof; comprises the amino acid sequence of SEQ ID NO: 30 and a N-terminal and/or C-terminal His-tag and/or HQ-tag; or is a fragment thereof having galactanase activity and having at least 90% such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% of the length of the mature polypeptide. In another embodiment, the polypeptide comprises or consists of the mature polypeptide of SEQ ID NO: 30. In another embodiment, the polypep- tide comprises or consists of amino acids 1 to 307 of SEQ ID NO: 30. In an embodiment, the polypeptide has been isolated.

In a continuation of the eighth aspect, the invention further relates to polypeptides having galac- tanase activity having a sequence identity to SEQ ID NO: 31 of at least 95%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 31 of at least 96%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 31 of at least 97%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 31 of at least 98%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 31 of at least 99%. In an embodiment, the polypeptide has a sequence identity to SEQ ID NO: 31 of at least 99.5%.

In one embodiment, the polypeptides differ by up to 16 amino acids, e.g., between 1 and 16 amino acids, such as 1-15, 1-14, 1-13, 1-12, 1-1 1 , 1-10, 1-9, 1-8, 1-7, 1-6 or 1-5 amino acids, or 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15 or 16 amino acids from SEQ ID NO: 31. In one embodiment, the polypeptides differ by up to 50 amino acids, e.g., between 1 and 16 amino ac- ids, such as 1-15, 1-14, 1-13, 1-12, 1-1 1 , 1-10, 1-9, 1-8, 1-7, 1-6 or 1-5 amino acids, or 1 , 2, 3,

4, 5, 6, 7, 8, 9, 10, 11 , 12, 13 ,14, 15 or 16 amino acids from SEQ ID NO: 32.

In one embodiment, the polypeptide preferably comprises or consists of the amino acid se- quence of SEQ ID NO: 31 or an allelic variant thereof; comprises the amino acid sequence of SEQ ID NO: 31 and a N-terminal and/or C-terminal His-tag and/or HQ-tag such as SEQ ID NO: 32; or is a fragment thereof having galactanase activity and having at least 90% such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% of the length of the mature polypeptide. In another embodiment, the poly- peptide comprises or consists of amino acids 1 to 307 of SEQ ID NO: 31. In another embodi- merit, the polypeptide comprises or consists of amino acids 1 to 315 of SEQ ID NO: 32. In an embodiment, the polypeptide has been isolated.

In a continuation of the eighth aspect, the invention relates to a polypeptide having galactanase activity encoded by a polynucleotide that hybridizes under high or very high stringency condi- tions with (i) the mature polypeptide coding sequence of SEQ ID NO: 29, (ii) the cDNA se- quence thereof, or (iii) the full-length complement of (i) or (ii) (Sambrook et al., supra). In an embodiment, the polypeptide has been isolated.

In a continuation of the eighth aspect, the invention relates to a polypeptide having galactanase activity encoded by a polynucleotide having a sequence identity to the mature polypeptide cod- ing sequence of SEQ ID NO: 29 or the cDNA sequence thereof of at least 95%, e.g. at least 96%, at least 97%, at least 98%, at least 99%, or 100%. In a further embodiment, the polypep- tide has been isolated.

In a continuation of the eighth aspect, the invention relates to variants of SEQ ID NO: 31 having galactanase activity comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof at one or more (e.g., several) positions. In an embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in SEQ ID NO: 31 is not more than 16, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15 or 16. In another embodiment, the number of posi- tions comprising one or more amino acid substitutions, and/or one or more amino acid dele- tions, and/or one or more amino acid insertions or any combination thereof in SEQ ID NO: 31 is between 1 and 15, such as 1-14, 1-13, 1-12, 1-11 , 1-10, 1-9, 1-8, 1-7, 1-6, or 1-5 positions. In an embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any com- bination thereof in SEQ ID NO: 31 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10. In an- other embodiment, the number of substitutions, deletions, and/or insertions in SEQ ID NO: 31 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further embodiment, the number of substitutions in SEQ ID NO: 31 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a fur- ther embodiment, the number of conservative substitutions in SEQ ID NO: 31 is not more than 10, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9 or 10.

In an embodiment, the polypeptide of the eighth aspect comprises the motif GV[T/M]PD[W/M]VQ[I/V]GNE (SEQ ID NO: 26) and/or the motif WADP[A/G]xQxKPxAW (SEQ ID NO: 27). Sources of Polypeptides Having Galactanase Activity

A polypeptide having galactanase activity of the present invention may be obtained from micro- organisms of any genus. For purposes of the present invention, the term“obtained from” as used herein in connection with a given source shall mean that the polypeptide encoded by a polynucleotide is produced by the source or by a strain in which the polynucleotide from the source has been inserted. In one embodiment, the polypeptide obtained from a given source is secreted extracellularly.

The polypeptide having galactanase activity is a bacterial polypeptide. In one embodiment, the polypeptide having galactanase activity is from the genus Cohnella or from the species Cohnel- la sp-60555, Cohnella xylanilytica or Cohnella laeviribosi. In another embodiment, the polypep- tide having galactanase activity is from the genus Paenibacillus or from the species Paenibacil- lus sp-18179, Paenibacillus peoriae or Paenibacillus xylanexedens. In another embodiment, the polypeptide having galactanase activity is from the genus Xanthomonas or from the species Xanthomonas cynarae.

It will be understood that for the aforementioned species, the invention encompasses both the perfect and imperfect states, and other taxonomic equivalents, e.g., anamorphs, regardless of the species name by which they are known. Those skilled in the art will readily recognize the identity of appropriate equivalents.

Strains of these species are readily accessible to the public in a number of culture collections, such as the American Type Culture Collection (ATCC), Deutsche Sammlung von Mikroorganis- men und Zellkulturen GmbH (DSMZ), Centraalbureau Voor Schimmelcultures (CBS), and Agri- cultural Research Service Patent Culture Collection, Northern Regional Research Center (NRRL).

The polypeptide may be identified and obtained from other sources including microorganisms isolated from nature (e.g., soil, composts, water, etc.) or DNA samples obtained directly from natural materials (e.g., soil, composts, water, etc.) using the above-mentioned probes. Tech- niques for isolating microorganisms and DNA directly from natural habitats are well known in the art. A polynucleotide encoding the polypeptide may then be obtained by similarly screening a genomic DNA or cDNA library of another microorganism or mixed DNA sample. Once a polynu- cleotide encoding a polypeptide has been detected with the probe(s), the polynucleotide can be isolated or cloned by utilizing techniques that are known to those of ordinary skill in the art (see, e.g., Sambrook et al., 1989, supra). Polynucleotides

The present invention also relates to isolated polynucleotides encoding a polypeptide of the present invention.

The techniques used to isolate or clone a polynucleotide are known in the art and include isola- tion from genomic DNA or cDNA, or a combination thereof. The cloning of the polynucleotides from genomic DNA can be effected, e.g., by using the well-known polymerase chain reaction (PCR) or antibody screening of expression libraries to detect cloned DNA fragments with shared structural features. See, e.g., Innis et al, 1990, PCR: A Guide to Methods and Application, Aca demic Press, New York. Other nucleic acid amplification procedures such as ligase chain reac- tion (LCR), ligation activated transcription (LAT) and polynucleotide-based amplification (NAS- BA) may be used. The polynucleotides may be cloned from a strain of Bacillus, or a related or- ganism and thus, for example, may be an allelic or species variant of the polypeptide encoding region of the polynucleotide.

Modification of a polynucleotide encoding a polypeptide of the present invention may be neces- sary for synthesizing polypeptides substantially similar to the polypeptide. The term“substantial- ly similar” to the polypeptide refers to non-naturally occurring forms of the polypeptide.

Nucleic Acid Constructs

The present invention also relates to nucleic acid constructs comprising a polynucleotide of the present invention operably linked to one or more control sequences that direct the expression of the coding sequence in a suitable host cell under conditions compatible with the control se- quences.

The polynucleotide may be manipulated in a variety of ways to provide for expression of the polypeptide. Manipulation of the polynucleotide prior to its insertion into a vector may be desira- ble or necessary depending on the expression vector. The techniques for modifying polynucleo- tides utilizing recombinant DNA methods are well known in the art.

The control sequence may be a promoter, a polynucleotide that is recognized by a host cell for expression of a polynucleotide encoding a polypeptide of the present invention. The promoter contains transcriptional control sequences that mediate the expression of the polypeptide. The promoter may be any polynucleotide that shows transcriptional activity in the host cell including mutant, truncated, and hybrid promoters, and may be obtained from genes encoding extracellu- lar or intracellular polypeptides either homologous or heterologous to the host cell.

Examples of suitable promoters for directing transcription of the nucleic acid constructs of the present invention in a bacterial host cell are the promoters obtained from the Bacillus amyloliq- uefaciens alpha-amylase gene ( amyQ ), Bacillus licheniformis alpha-amylase gene ( amyL ), Ba cillus licheniformis penicillinase gene ( penP ), Bacillus stearothermophilus maltogenic amylase gene ( amyM ), Bacillus subtilis levansucrase gene (sacS), Bacillus subtilis xylA and xylB genes, Bacillus thuringiensis crylllA gene (Agaisse and Lereclus, 1994, Molecular Microbiology 13: 97- 107), E. coli lac operon, E. coli trc promoter (Egon et al., 1988, Gene 69: 301-315), Streptomy- ces coelicolor agarase gene ( dagA ), and prokaryotic beta-lactamase gene (Villa-Kamaroff et al., 1978, Proc. Natl. Acad. Sci. USA 75: 3727-3731 ), as well as the tac promoter (DeBoer et al., 1983, Proc. Natl. Acad. Sci. USA 80: 21-25). Further promoters are described in "Useful pro- teins from recombinant bacteria" in Gilbert et al., 1980, Scientific American 242: 74-94; and in Sambrook et al., 1989, supra. Examples of tandem promoters are disclosed in WO 99/43835.

Examples of suitable promoters for directing transcription of the nucleic acid constructs of the present invention in a filamentous fungal host cell are promoters obtained from the genes for Aspergillus nidulans acetamidase, Aspergillus niger neutral alpha-amylase, Aspergillus niger acid stable alpha-amylase, Aspergillus niger or Aspergillus awamori glucoamylase ( glaA ), As pergillus oryzae TAKA amylase, Aspergillus oryzae alkaline protease, Aspergillus oryzae triose phosphate isomerase, Fusarium oxysporum trypsin-like protease (WO 96/00787), Fusarium venenatum amyloglucosidase (WO 00/56900), Fusarium venenatum Daria (WO 00/56900), Fusarium venenatum Quinn (WO 00/56900), Rhizomucor miehei lipase, Rhizomucor miehei as partic proteinase, Trichoderma reesei beta-glucosidase, Trichoderma reesei cellobiohydrolase I, Trichoderma reesei cellobiohydrolase II, Trichoderma reesei endoglucanase I, Trichoderma reesei endoglucanase II, Trichoderma reesei endoglucanase III, Trichoderma reesei endoglu- canase V, Trichoderma reesei galactanase I, Trichoderma reesei galactanase II, Trichoderma reesei galactanase III, Trichoderma reesei beta-xylosidase, and Trichoderma reesei translation elongation factor, as well as the NA2-tpi promoter (a modified promoter from an Aspergillus neu- tral alpha-amylase gene in which the untranslated leader has been replaced by an untranslated leader from an Aspergillus triose phosphate isomerase gene; non-limiting examples include modified promoters from an Aspergillus niger neutral alpha-amylase gene in which the untrans- lated leader has been replaced by an untranslated leader from an Aspergillus nidulans or As pergillus oryzae triose phosphate isomerase gene); and mutant, truncated, and hybrid promot- ers thereof. Other promoters are described in U.S. Patent No. 6,011 ,147.

In a yeast host, useful promoters are obtained from the genes for Saccharomyces cerevisiae enolase (ENO-1 ), Saccharomyces cerevisiae galactokinase (GAL1 ), Saccharomyces cerevisiae alcohol dehydrogenase/glyceraldehyde-3-phosphate dehydrogenase (ADH1 , ADH2/GAP), Sac charomyces cerevisiae triose phosphate isomerase (TPI), Saccharomyces cerevisiae metal- lothionein (CUP1 ), and Saccharomyces cerevisiae 3-phosphoglycerate kinase. Other useful promoters for yeast host cells are described by Romanos et al., 1992, Yeast 8: 423-488.

The control sequence may also be a transcription terminator, which is recognized by a host cell to terminate transcription. The terminator is operably linked to the 3’-terminus of the polynucleo- tide encoding the polypeptide. Any terminator that is functional in the host cell may be used in the present invention.

Preferred terminators for bacterial host cells are obtained from the genes for Bacillus clausii al kaline protease ( aprH ), Bacillus licheniformis alpha-amylase ( amyL ), and Escherichia coli ribo- somal RNA ( rrnB ).

Preferred terminators for filamentous fungal host cells are obtained from the genes for Aspergil lus nidulans acetamidase, Aspergillus nidulans anthranilate synthase, Aspergillus niger glu- coamylase, Aspergillus niger alpha-glucosidase, Aspergillus oryzae TAKA amylase, Fusarium oxysporum trypsin-like protease, Trichoderma reesei beta-glucosidase, Trichoderma reesei cel- lobiohydrolase I, Trichoderma reesei cellobiohydrolase II, Trichoderma reesei endoglucanase I, Trichoderma reesei endoglucanase II, Trichoderma reesei endoglucanase III, Trichoderma reesei endoglucanase V, Trichoderma reesei galactanase I, Trichoderma reesei galactanase II, Trichoderma reesei galactanase III, Trichoderma reesei beta-xylosidase, and Trichoderma reesei translation elongation factor.

Preferred terminators for yeast host cells are obtained from the genes for Saccharomyces cere- visiae enolase, Saccharomyces cerevisiae cytochrome C (CYC1 ), and Saccharomyces cere- visiae glyceraldehyde-3-phosphate dehydrogenase. Other useful terminators for yeast host cells are described by Romanos et al., 1992, supra.

The control sequence may also be an mRNA stabilizer region downstream of a promoter and upstream of the coding sequence of a gene which increases expression of the gene.

Examples of suitable mRNA stabilizer regions are obtained from a Bacillus thuringiensis crylllA gene (WO 94/25612) and a Bacillus subtilis SP82 gene (Hue ef a/., 1995, Journal of Bacteriolo gy 177: 3465-3471 ).

The control sequence may also be a leader, a nontranslated region of an mRNA that is im- portant for translation by the host cell. The leader is operably linked to the 5’-terminus of the polynucleotide encoding the polypeptide. Any leader that is functional in the host cell may be used.

Preferred leaders for filamentous fungal host cells are obtained from the genes for Aspergillus oryzae TAKA amylase and Aspergillus nidulans triose phosphate isomerase.

Suitable leaders for yeast host cells are obtained from the genes for Saccharomyces cerevisiae enolase (ENO-1 ), Saccharomyces cerevisiae 3-phosphoglycerate kinase, Saccharomyces cerevisiae alpha-factor, and Saccharomyces cerevisiae alcohol dehydrogenase/glyceraldehyde- 3-phosphate dehydrogenase (ADH2/GAP).

The control sequence may also be a polyadenylation sequence, a sequence operably linked to the 3’-terminus of the polynucleotide and, when transcribed, is recognized by the host cell as a signal to add polyadenosine residues to transcribed mRNA. Any polyadenylation sequence that is functional in the host cell may be used.

Preferred polyadenylation sequences for filamentous fungal host cells are obtained from the genes for Aspergillus nidulans anthranilate synthase, Aspergillus niger glucoamylase, Aspergil lus niger alpha-glucosidase Aspergillus oryzae TAKA amylase, and Fusarium oxysporum tryp- sin-like protease.

Useful polyadenylation sequences for yeast host cells are described by Guo and Sherman, 1995, Mol. Cellular Biol. 15: 5983-5990.

The control sequence may also be a signal peptide coding region that encodes a signal peptide linked to the N-terminus of a polypeptide and directs the polypeptide into the cell’s secretory pathway. The 5’-end of the coding sequence of the polynucleotide may inherently contain a sig- nal peptide coding sequence naturally linked in translation reading frame with the segment of the coding sequence that encodes the polypeptide. Alternatively, the 5’-end of the coding se- quence may contain a signal peptide coding sequence that is foreign to the coding sequence. A foreign signal peptide coding sequence may be required where the coding sequence does not naturally contain a signal peptide coding sequence. Alternatively, a foreign signal peptide cod- ing sequence may simply replace the natural signal peptide coding sequence in order to en- hance secretion of the polypeptide. However, any signal peptide coding sequence that directs the expressed polypeptide into the secretory pathway of a host cell may be used.

Effective signal peptide coding sequences for bacterial host cells are the signal peptide coding sequences obtained from the genes for Bacillus NCIB 1 1837 maltogenic amylase, Bacillus li- cheniformis subtilisin, Bacillus licheniformis beta-lactamase, Bacillus stearothermophilus alpha- amylase, Bacillus stearothermophilus neutral proteases ( nprT , nprS, nprM), and Bacillus subtilis prsA. Further signal peptides are described by Simonen and Palva, 1993, Microbiological Re views 57: 109-137.

Effective signal peptide coding sequences for filamentous fungal host cells are the signal pep- tide coding sequences obtained from the genes for Aspergillus niger neutral amylase, Aspergil lus niger glucoamylase, Aspergillus oryzae TAKA amylase, Humicola insolens cellulase, Humi- cola insolens endoglucanase V, Humicola lanuginosa lipase, and Rhizomucor miehei aspartic proteinase.

Useful signal peptides for yeast host cells are obtained from the genes for Saccharomyces cerevisiae alpha-factor and Saccharomyces cerevisiae invertase. Other useful signal peptide coding sequences are described by Romanos et a!., 1992, supra.

The control sequence may also be a propeptide coding sequence that encodes a propeptide positioned at the N-terminus of a polypeptide. The resultant polypeptide is known as a proen- zyme or propolypeptide (or a zymogen in some cases). A propolypeptide is generally inactive and can be converted to an active polypeptide by catalytic or autocatalytic cleavage of the pro- peptide from the propolypeptide. The propeptide coding sequence may be obtained from the genes for Bacillus subtilis alkaline protease (aprE), Bacillus subtilis neutral protease ( nprT ), My- celiophthora thermophila laccase (WO 95/33836), Rhizomucor miehei aspartic proteinase, and Saccharomyces cerevisiae alpha-factor.

Where both signal peptide and propeptide sequences are present, the propeptide sequence is positioned next to the N-terminus of a polypeptide and the signal peptide sequence is posi- tioned next to the N-terminus of the propeptide sequence.

It may also be desirable to add regulatory sequences that regulate expression of the polypep- tide relative to the growth of the host cell. Examples of regulatory sequences are those that cause expression of the gene to be turned on or off in response to a chemical or physical stimu- lus, including the presence of a regulatory compound. Regulatory sequences in prokaryotic sys- tems include the lac, tac, and trp operator systems. In yeast, the ADH2 system or GAL1 system may be used. In filamentous fungi, the Aspergillus niger glucoamylase promoter, Aspergillus oryzae TAKA alpha-amylase promoter, and Aspergillus oryzae glucoamylase promoter, Tricho- derma reesei cellobiohydrolase I promoter, and Trichoderma reesei cellobiohydrolase II pro- moter may be used. Other examples of regulatory sequences are those that allow for gene am- plification. In eukaryotic systems, these regulatory sequences include the dihydrofolate reduc- tase gene that is amplified in the presence of methotrexate, and the metallothionein genes that are amplified with heavy metals. In these cases, the polynucleotide encoding the polypeptide would be operably linked to the regulatory sequence.

Expression Vectors

The present invention also relates to recombinant expression vectors comprising a polynucleo- tide of the present invention, a promoter, and transcriptional and translational stop signals. The various nucleotide and control sequences may be joined together to produce a recombinant ex- pression vector that may include one or more convenient restriction sites to allow for insertion or substitution of the polynucleotide encoding the polypeptide at such sites. Alternatively, the poly- nucleotide may be expressed by inserting the polynucleotide or a nucleic acid construct corn- prising the polynucleotide into an appropriate vector for expression. In creating the expression vector, the coding sequence is located in the vector so that the coding sequence is operably linked with the appropriate control sequences for expression.

The recombinant expression vector may be any vector (e.g., a plasmid or virus) that can be conveniently subjected to recombinant DNA procedures and can bring about expression of the polynucleotide. The choice of the vector will typically depend on the compatibility of the vector with the host cell into which the vector is to be introduced. The vector may be a linear or closed circular plasmid. The vector may be an autonomously replicating vector, i.e., a vector that exists as an ex- trachromosomal entity, the replication of which is independent of chromosomal replication, e.g., a plasmid, an extrachromosomal element, a minichromosome, or an artificial chromosome. The vector may contain any means for assuring self-replication. Alternatively, the vector may be one that, when introduced into the host cell, is integrated into the genome and replicated together with the chromosome(s) into which it has been integrated. Furthermore, a single vector or plas- mid or two or more vectors or plasmids that together contain the total DNA to be introduced into the genome of the host cell, or a transposon, may be used.

The vector preferably contains one or more selectable markers that permit easy selection of transformed, transfected, transduced, or the like cells. A selectable marker is a gene the product of which provides for biocide or viral resistance, resistance to heavy metals, prototrophy to aux- otrophs, and the like.

Examples of bacterial selectable markers are Bacillus licheniformis or Bacillus subtilis dal genes, or markers that confer antibiotic resistance such as ampicillin, chloramphenicol, kana- mycin, neomycin, spectinomycin, or tetracycline resistance. Suitable markers for yeast host cells include, but are not limited to, ADE2, HIS3, LEU2, LYS2, MET3, TRP1 , and URA3. Se- lectable markers for use in a filamentous fungal host cell include, but are not limited to, adeA (phosphoribosylaminoimidazole-succinocarboxamide synthase), adeB (phosphoribosyl- aminoimidazole synthase), amdS (acetamidase), argB (ornithine carbamoyltransferase), bar (phosphinothricin acetyltransferase), hph (hygromycin phosphotransferase), niaD (nitrate reduc- tase), pyrG (orotidine-5’-phosphate decarboxylase), sC (sulfate adenyltransferase), and trpC (anthranilate synthase), as well as equivalents thereof. Preferred for use in an Aspergillus cell are Aspergillus nidulans or Aspergillus oryzae amdS and pyrG genes and a Streptomyces hy- groscopicus bar gene. Preferred for use in a Trichoderma cell are adeA, adeB, amdS, hph, and pyrG genes.

The selectable marker may be a dual selectable marker system as described in WO 2010/039889. In one aspect, the dual selectable marker is an hph-tk dual selectable marker system.

The vector preferably contains an element(s) that permits integration of the vector into the host cell's genome or autonomous replication of the vector in the cell independent of the genome.

For integration into the host cell genome, the vector may rely on the polynucleotide’s sequence encoding the polypeptide or any other element of the vector for integration into the genome by homologous or non-homologous recombination. Alternatively, the vector may contain additional polynucleotides for directing integration by homologous recombination into the genome of the host cell at a precise location(s) in the chromosome(s). To increase the likelihood of integration at a precise location, the integrational elements should contain a sufficient number of nucleic acids, such as 100 to 10,000 base pairs, 400 to 10,000 base pairs, and 800 to 10,000 base pairs, which have a high degree of sequence identity to the corresponding target sequence to enhance the probability of homologous recombination. The integrational elements may be any sequence that is homologous with the target sequence in the genome of the host cell. Further- more, the integrational elements may be non-encoding or encoding polynucleotides. On the other hand, the vector may be integrated into the genome of the host cell by non-homologous recombination.

For autonomous replication, the vector may further comprise an origin of replication enabling the vector to replicate autonomously in the host cell in question. The origin of replication may be any plasmid replicator mediating autonomous replication that functions in a cell. The term“origin of replication” or“plasmid replicator” means a polynucleotide that enables a plasmid or vector to replicate in vivo.

Examples of bacterial origins of replication are the origins of replication of plasmids pBR322, pUC19, pACYC177, and pACYC184 permitting replication in E. coli, and pUB1 10, pE194, pTA1060, and rAMb1 permitting replication in Bacillus.

Examples of origins of replication for use in a yeast host cell are the 2 micron origin of replica- tion, ARS1 , ARS4, the combination of ARS1 and CEN3, and the combination of ARS4 and CEN6.

Examples of origins of replication useful in a filamentous fungal cell are AMA1 and ANSI (Gems et al., 1991 , Gene 98: 61-67; Cullen et al., 1987, Nucleic Acids Res. 15: 9163-9175; WO 00/24883). Isolation of the AMA1 gene and construction of plasmids or vectors comprising the gene can be accomplished according to the methods disclosed in WO 00/24883.

More than one copy of a polynucleotide of the present invention may be inserted into a host cell to increase production of a polypeptide. An increase in the copy number of the polynucleotide can be obtained by integrating at least one additional copy of the sequence into the host cell genome or by including an amplifiable selectable marker gene with the polynucleotide where cells containing amplified copies of the selectable marker gene, and thereby additional copies of the polynucleotide, can be selected for by cultivating the cells in the presence of the appropriate selectable agent.

The procedures used to ligate the elements described above to construct the recombinant ex- pression vectors of the present invention are well known to one skilled in the art (see, e.g., Sambrook et al., 1989, supra).

Host Cells

The present invention also relates to recombinant host cells, comprising a polynucleotide of the present invention operably linked to one or more control sequences that direct the production of a polypeptide of the present invention. A construct or vector comprising a polynucleotide is in- traduced into a host cell so that the construct or vector is maintained as a chromosomal inte grant or as a self-replicating extra-chromosomal vector as described earlier. The term "host cell" encompasses any progeny of a parent cell that is not identical to the parent cell due to muta- tions that occur during replication. The choice of a host cell will to a large extent depend upon the gene encoding the polypeptide and its source.

The host cell may be any cell useful in the recombinant production of a polypeptide of the pre- sent invention, e.g., a prokaryote or a eukaryote.

The prokaryotic host cell may be any Gram-positive or Gram-negative bacterium. Gram-positive bacteria include, but are not limited to, Bacillus, Clostridium, Enterococcus, Geobacillus, Lacto bacillus, Lactococcus, Oceanobacillus, Staphylococcus, Streptococcus, and Streptomyces. Gram-negative bacteria include, but are not limited to, Campylobacter, E. coli, Flavobacterium, Fusobacterium, Helicobacter, llyobacter, Neisseria, Pseudomonas, Salmonella, and Ureaplas- ma.

The bacterial host cell may be any Bacillus cell including, but not limited to, Bacillus al- kalophilus, Bacillus amyloliquefaciens, Bacillus brevis, Bacillus circulans, Bacillus clausii, Bacil- lus coagulans, Bacillus firmus, Bacillus lautus, Bacillus lentus, Bacillus licheniformis, Bacillus megaterium, Bacillus pumilus, Bacillus stearothermophilus, Bacillus subtilis, and Bacillus thurin- giensis cells.

The bacterial host cell may also be any Streptococcus cell including, but not limited to, Strepto- coccus equisimilis, Streptococcus pyogenes, Streptococcus uberis, and Streptococcus equi subsp. Zooepidemicus cells.

The bacterial host cell may also be any Streptomyces cell including, but not limited to, Strepto- myces achromogenes, Streptomyces avermitilis, Streptomyces coelicolor, Streptomyces griseus, and Streptomyces lividans cells.

The introduction of DNA into a Bacillus cell may be effected by protoplast transformation (see, e.g., Chang and Cohen, 1979, Mol. Gen. Genet. 168: 11 1-1 15), competent cell transformation (see, e.g., Young and Spizizen, 1961 , J. Bacteriol. 81 : 823-829, or Dubnau and Davidoff- Abelson, 1971 , J. Mol. Biol. 56: 209-221 ), electroporation (see, e.g., Shigekawa and Dower,

1988, Biotechniques 6: 742-751 ), or conjugation (see, e.g., Koehler and Thorne, 1987, J. Bacte riol. 169: 5271-5278). The introduction of DNA into an E. coli cell may be effected by protoplast transformation (see, e.g., Hanahan, 1983, J. Mol. Biol. 166: 557-580) or electroporation (see, e.g., Dower et a!., 1988, Nucleic Acids Res. 16: 6127-6145). The introduction of DNA into a Streptomyces cell may be effected by protoplast transformation, electroporation (see, e.g., Gong et al., 2004, Folia Microbiol. {Praha) 49: 399-405), conjugation (see, e.g., Mazodier et al.,

1989, J. Bacteriol. 171 : 3583-3585), or transduction (see, e.g., Burke et al., 2001 , Proc. Natl. Acad. Sci. USA 98: 6289-6294). The introduction of DNA into a Pseudomonas cell may be ef- fected by electroporation (see, e.g., Choi et al., 2006, J. Microbiol. Methods 64: 391-397) or conjugation (see, e.g., Pinedo and Smets, 2005, Appl. Environ. Microbiol. 71 : 51-57). The intro- duction of DNA into a Streptococcus cell may be effected by natural competence (see, e.g., Per- ry and Kuramitsu, 1981 , Infect. Immun. 32: 1295-1297), protoplast transformation (see, e.g., Catt and Jollick, 1991 , Microbios 68: 189-207), electroporation (see, e.g., Buckley et al., 1999, Appl. Environ. Microbiol. 65: 3800-3804), or conjugation (see, e.g., Clewell, 1981 , Microbiol. Rev. 45: 409-436). However, any method known in the art for introducing DNA into a host cell can be used.

The host cell may also be a eukaryote, such as a mammalian, insect, plant, or fungal cell.

The host cell may be a fungal cell.“Fungi” as used herein includes the phyla Ascomycota, Ba- sidiomycota, Chytridiomycota, and Zygomycota as well as the Oomycota and all mitosporic fun- gi (as defined by Hawksworth et al., In, Ainsworth and Bisby’s Dictionary of The Fungi, 8th edi- tion, 1995, CAB International, University Press, Cambridge, UK).

The fungal host cell may be a yeast cell.“Yeast” as used herein includes ascosporogenous yeast (Endomycetales), basidiosporogenous yeast, and yeast belonging to the Fungi Imperfecti (Blastomycetes). Since the classification of yeast may change in the future, for the purposes of this invention, yeast shall be defined as described in Biology and Activities of Yeast (Skinner, Passmore, and Davenport, editors, Soc. App. Bacteriol. Symposium Series No. 9, 1980).

The yeast host cell may be a Candida, Hansenula, Kluyveromyces, Pichia, Saccharomyces, Schizosaccharomyces, or Yarrowia cell, such as a Kluyveromyces lactis, Saccharomyces carls- bergensis, Saccharomyces cerevisiae, Saccharomyces diastaticus, Saccharomyces douglasii, Saccharomyces kluyveri, Saccharomyces norbensis, Saccharomyces oviformis, or Yarrowia lipolytica cell.

The fungal host cell may be a filamentous fungal cell.“Filamentous fungi” include all filamentous forms of the subdivision Eumycota and Oomycota (as defined by Hawksworth et al., 1995, su pra). The filamentous fungi are generally characterized by a mycelial wall composed of chitin, cellulose, glucan, chitosan, mannan, and other complex polysaccharides. Vegetative growth is by hyphal elongation and carbon catabolism is obligately aerobic. In contrast, vegetative growth by yeasts such as Saccharomyces cerevisiae is by budding of a unicellular thallus and carbon catabolism may be fermentative.

The filamentous fungal host cell may be an Acremonium, Aspergillus, Aureobasidium, Bjerkan- dera, Ceriporiopsis, Chrysosporium, Coprinus, Coriolus, Cryptococcus, Filibasidium, Fusarium, Humicola, Magnaporthe, Mucor, Myceliophthora, Neocallimastix, Neurospora, Paecilomyces, Penicillium, Phanerochaete, Phlebia, Piromyces, Pleurotus, Schizophyllum, Talaromyces, Thermoascus, Thielavia, Tolypocladium, Trametes, or Trichoderma cell. For example, the filamentous fungal host cell may be an Aspergillus awamori, Aspergillus foeti- dus, Aspergillus fumigatus, Aspergillus japonicus, Aspergillus nidulans, Aspergillus niger, As- pergillus oryzae, Bjerkandera adusta, Ceriporiopsis aneirina, Ceriporiopsis caregiea, Ceripori- opsis gilvescens, Ceriporiopsis pannocinta, Ceriporiopsis rivulosa, Ceriporiopsis subrufa, Ceri- poriopsis subvermispora, Chrysosporium inops, Chrysosporium keratinophilum, Chrysosporium lucknowense, Chrysosporium merdarium, Chrysosporium pannicola, Chrysosporium queens- landicum, Chrysosporium tropicum, Chrysosporium zonatum, Coprinus cinereus, Coriolus hirsu- tus, Fusarium bactridioides, Fusarium cerealis, Fusarium crookwellense, Fusarium culmorum, Fusarium graminearum, Fusarium graminum, Fusarium heterosporum, Fusarium negundi, Fusarium oxysporum, Fusarium reticulatum, Fusarium roseum, Fusarium sambucinum, Fusari- um sarcochroum, Fusarium sporotrichioides, Fusarium sulphureum, Fusarium torulosum, Fusarium trichothecioides, Fusarium venenatum, Humicola insolens, Humicola lanuginosa, Mu- cor miehei, Myceliophthora thermophila, Neurospora crassa, Penicillium purpurogenum, Phan- erochaete chrysosporium, Phlebia radiata, Pleurotus eryngii, Thielavia terrestris, Trametes vil- losa, Trametes versicolor, Trichoderma harzianum, Trichoderma koningii, Trichoderma longi- brachiatum, Trichoderma reesei, or Trichoderma viride cell.

Fungal cells may be transformed by a process involving protoplast formation, transformation of the protoplasts, and regeneration of the cell wall in a manner known per se. Suitable procedures for transformation of Aspergillus and Trichoderma host cells are described in EP 238023, Yelton et a!., 1984, Proc. Natl. Acad. Sci. USA 81 : 1470-1474, and Christensen et a!., 1988, Bio/Technology 6: 1419-1422. Suitable methods for transforming Fusarium species are de- scribed by Malardier et a!., 1989, Gene 78: 147-156, and WO 96/00787. Yeast may be trans- formed using the procedures described by Becker and Guarente, In Abelson, J.N. and Simon, M.I., editors, Guide to Yeast Genetics and Molecular Biology, Methods in Enzymology, Volume 194, pp 182-187, Academic Press, Inc., New York; Ito et a!., 1983, J. Bacteriol. 153: 163; and Hinnen et a!., 1978, Proc. Natl. Acad. Sci. USA 75: 1920.

Methods of Production

The present invention also relates to methods of producing a polypeptide of the present inven- tion, comprising (a) cultivating a cell, which in its wild-type form produces the polypeptide, under conditions conducive for production of the polypeptide; and optionally, (b) recovering the poly- peptide. In one aspect, the cell is a Cohnella cell. In another aspect, the cell is a Cohnella sp- 60555 cell. In a further aspect, the cell is a Cohnella xylanilytica cell. In a further aspect, the cell is a Cohnella laeviribosi cell. In one aspect, the cell is a Paenibacillus cell. In another aspect, the cell is a Paenibacillus sp-18179 cell. In a further aspect, the cell is a Paenibacillus peoriae cell. In another aspect, the cell is a Paenibacillus xylanexedens cell. In one aspect, the cell is a Xanthomonas cell. In another aspect, the cell is a Xanthomonas cynarae cell. The present invention also relates to methods of producing a polypeptide of the present inven- tion, comprising (a) cultivating a recombinant host cell of the present invention under conditions conducive for production of the polypeptide; and optionally, (b) recovering the polypeptide.

The host cells are cultivated in a nutrient medium suitable for production of the polypeptide us- ing methods known in the art. For example, the cells may be cultivated by shake flask cultiva- tion, or small-scale or large-scale fermentation (including continuous, batch, fed-batch, or solid state fermentations) in laboratory or industrial fermentors in a suitable medium and under condi- tions allowing the polypeptide to be expressed and/or isolated. The cultivation takes place in a suitable nutrient medium comprising carbon and nitrogen sources and inorganic salts, using procedures known in the art. Suitable media are available from commercial suppliers or may be prepared according to published compositions (e.g., in catalogues of the American Type Culture Collection). If the polypeptide is secreted into the nutrient medium, the polypeptide can be re- covered directly from the medium. If the polypeptide is not secreted, it can be recovered from cell lysates.

The polypeptide may be detected using methods known in the art that are specific for the poly- peptides. These detection methods include, but are not limited to, use of specific antibodies, formation of an enzyme product, or disappearance of an enzyme substrate. For example, an enzyme assay may be used to determine the activity of the polypeptide.

The polypeptide may be recovered using methods known in the art. For example, the polypep- tide may be recovered from the nutrient medium by conventional procedures including, but not limited to, collection, centrifugation, filtration, extraction, spray-drying, evaporation, or precipita- tion. In one aspect, a fermentation broth comprising the polypeptide is recovered.

The polypeptide may be purified by a variety of procedures known in the art including, but not limited to, chromatography (e.g., ion exchange, affinity, hydrophobic, chromatofocusing, and size exclusion), electrophoretic procedures (e.g., preparative isoelectric focusing), differential solubility (e.g., ammonium sulfate precipitation), SDS-PAGE, or extraction (see, e.g., Protein Purification, Janson and Ryden, editors, VCH Publishers, New York, 1989) to obtain substantial- ly pure polypeptides.

In an alternative aspect, the polypeptide is not recovered, but rather a host cell of the present invention expressing the polypeptide is used as a source of the polypeptide.

Production in plants

The present invention also relates to isolated plants, e.g., a transgenic plant, plant part, or plant cell, comprising a polynucleotide of the present invention so as to express and produce a poly- peptide or domain in recoverable quantities. The polypeptide or domain may be recovered from the plant or plant part. Alternatively, the plant or plant part containing the polypeptide or domain may be used as such for improving the quality of a food or feed, e.g., improving nutritional val- ue, palatability, and rheological properties, or to destroy an antinutritive factor.

The transgenic plant can be dicotyledonous (a dicot) or monocotyledonous (a monocot). Exam- pies of monocot plants are grasses, such as meadow grass (blue grass, Poa ), forage grass such as Festuca, Lolium, temperate grass, such as Agrostis, and cereals, e.g., wheat, oats, rye, barley, rice, sorghum, and maize (corn).

Examples of dicot plants are tobacco, legumes, such as lupins, potato, sugar beet, pea, bean and soybean, and cruciferous plants (family Brassicaceae), such as cauliflower, rape seed, and the closely related model organism Arabidopsis thaliana.

Examples of plant parts are stem, callus, leaves, root, fruits, seeds, and tubers as well as the individual tissues comprising these parts, e.g., epidermis, mesophyll, parenchyme, vascular tis sues, meristems.

Plant cells and specific plant cell compartments, such as chloroplasts, apoplasts, mitochondria, vacuoles, peroxisomes and cytoplasm are also considered to be a plant part.

Also included within the scope of the present invention are the progeny of such plants, plant parts, and plant cells.

The transgenic plant or plant cell expressing the polypeptide or domain may be constructed in accordance with methods known in the art.

The present invention also relates to methods of producing a polypeptide or domain of the pre- sent invention comprising (a) cultivating a transgenic plant or a plant cell comprising a polynu- cleotide encoding the polypeptide or domain under conditions conducive for production of the polypeptide or domain; and (b) recovering the polypeptide or domain.

Fermentation Broth Formulations or Cell Compositions

The present invention also relates to a fermentation broth formulation or a cell composition comprising a polypeptide of the present invention. The fermentation broth product further corn- prises additional ingredients used in the fermentation process, such as, for example, cells (in- cluding, the host cells containing the gene encoding the polypeptide of the present invention which are used to produce the polypeptide of interest), cell debris, biomass, fermentation media and/or fermentation products. In some embodiments, the composition is a cell-killed whole broth containing organic acid(s), killed cells and/or cell debris, and culture medium.

The term "fermentation broth" as used herein refers to a preparation produced by cellular fer- mentation that undergoes no or minimal recovery and/or purification. For example, fermentation broths are produced when microbial cultures are grown to saturation, incubated under carbon- limiting conditions to allow protein synthesis (e.g., expression of enzymes by host cells) and se- cretion into cell culture medium. The fermentation broth can contain unfractionated or fraction- ated contents of the fermentation materials derived at the end of the fermentation. Typically, the fermentation broth is unfractionated and comprises the spent culture medium and cell debris present after the microbial cells (e.g., filamentous fungal cells) are removed, e.g., by centrifuga- tion. In some embodiments, the fermentation broth contains spent cell culture medium, extracel- lular enzymes, and viable and/or nonviable microbial cells.

In an embodiment, the fermentation broth formulation and cell compositions comprise a first or- ganic acid component comprising at least one 1-5 carbon organic acid and/or a salt thereof and a second organic acid component comprising at least one 6 or more carbon organic acid and/or a salt thereof. In a specific embodiment, the first organic acid component is acetic acid, formic acid, propionic acid, a salt thereof, or a mixture of two or more of the foregoing and the second organic acid component is benzoic acid, cyclohexanecarboxylic acid, 4-methylvaleric acid, phe- nylacetic acid, a salt thereof, or a mixture of two or more of the foregoing.

In one aspect, the composition contains an organic acid(s), and optionally further contains killed cells and/or cell debris. In one embodiment, the killed cells and/or cell debris are removed from a cell-killed whole broth to provide a composition that is free of these components.

The fermentation broth formulations or cell compositions may further comprise a preservative and/or anti-microbial (e.g., bacteriostatic) agent, including, but not limited to, sorbitol, sodium chloride, potassium sorbate, and others known in the art.

The cell-killed whole broth or composition may contain the unfractionated contents of the fer- mentation materials derived at the end of the fermentation. Typically, the cell-killed whole broth or composition contains the spent culture medium and cell debris present after the microbial cells (e.g., filamentous fungal cells) are grown to saturation, incubated under carbon-limiting conditions to allow protein synthesis. In some embodiments, the cell-killed whole broth or corn- position contains the spent cell culture medium, extracellular enzymes, and killed filamentous fungal cells. In some embodiments, the microbial cells present in the cell-killed whole broth or composition can be permeabilized and/or lysed using methods known in the art.

A whole broth or cell composition as described herein is typically a liquid, but may contain insol uble components, such as killed cells, cell debris, culture media components, and/or insoluble enzyme(s). In some embodiments, insoluble components may be removed to provide a clarified liquid composition.

The whole broth formulations and cell compositions of the present invention may be produced by a method described in WO 90/15861 or WO 2010/096673. Enzyme Compositions

Preferably, the compositions are enriched in the polypeptides of the invention. The term "en- riched" indicates that the galactanase activity of the composition has been increased, e.g., with an enrichment factor of at least 1.1 , such as at least 1.2, at least 1.3, at least 1.4, at least 1.5, at least 2.0, at least 3.0, at least 4.0, at least 5.0, at least 10. In an embodiment, the composition comprises the polypeptides of the invention and one or more formulating agents, as described in the‘formulating agent’ section below.

The present invention also relates to compositions comprising the polypeptide of the second aspect of the invention (SEQ ID NO: 3) having galactanase activity. In an embodiment, the composition further comprises one or more formulating agents.

The present invention also relates to compositions comprising the polypeptide of the third as- pect of the invention (SEQ ID NO: 7) having galactanase activity. In an embodiment, the corn- position further comprises one or more formulating agents.

The present invention also relates to compositions comprising the polypeptide of the fourth as- pect of the invention (SEQ ID NO: 11 ) having galactanase activity. In an embodiment, the corn- position further comprises one or more formulating agents.

The present invention also relates to compositions comprising the polypeptide of the fifth aspect of the invention (SEQ ID NO: 15) having galactanase activity. In an embodiment, the composi- tion further comprises one or more formulating agents.

The present invention also relates to compositions comprising the polypeptide of the sixth as- pect of the invention (SEQ ID NO: 19) having galactanase activity. In an embodiment, the corn- position further comprises one or more formulating agents.

The present invention also relates to compositions comprising the polypeptide of the seventh aspect of the invention (SEQ ID NO: 23) having galactanase activity. In an embodiment, the composition further comprises one or more formulating agents.

The present invention also relates to compositions comprising the polypeptide of the eighth as- pect of the invention (SEQ ID NO: 31 ) having galactanase activity. In an embodiment, the corn- position further comprises one or more formulating agents.

The compositions may comprise a polypeptide of the present invention as the major enzymatic component, e.g., a mono-component composition. Such a composition may further comprise a formulating agent, as described in the‘formulating agent’ section below. Alternatively, the corn- positions may comprise multiple enzymatic activities, such as one or more (e.g., several) en- zymes selected from the group consisting of phytase, galactanase, alpha-galactosidase, prote- ase, phospholipase, glucoronidase, lysophospholipase, amylase, beta-glucanase, beta- galactosidase, beta-xylosidase, endo-1 ,4-beta-galactanase acetyl xylan esterase, feruloyl es- terase, cellulase, cellobiohydrolase, beta-glycosidase, pullulanase, or any mixture thereof. It is at present contemplated that the galactanase is used in one or more of the following amounts (dosage ranges): 0.01-200; 0.05-100; 0.1-50; 0.2-20; 0.1-1 ; 0.2-2; 0.5-5; or 1-10 wherein all these ranges are mg galactanase protein per kg substrate (ppm).

Formulating agent

The enzyme of the invention may be formulated as a liquid or a solid. For a liquid formulation, the formulating agent may comprise a polyol (such as e.g. glycerol, ethylene glycol or propylene glycol), a salt (such as e.g. sodium chloride, sodium benzoate, potassium sorbate) or a sugar or sugar derivative (such as e.g. dextrin, glucose, sucrose, and sorbitol). Thus in one embodiment, the composition is a liquid composition comprising the polypeptide of the invention and one or more formulating agents selected from the list consisting of glycerol, ethylene glycol, 1 ,2- propylene glycol, 1 ,3-propylene glycol, sodium chloride, sodium benzoate, potassium sorbate, dextrin, glucose, sucrose, and sorbitol. The liquid formulation may be sprayed onto the feed af- ter it has been pelleted or may be added to drinking water given to the animals.

For a solid formulation, the formulation may be for example as a granule, spray dried powder or agglomerate. The formulating agent may comprise a salt (organic or inorganic zinc, sodium, potassium or calcium salts such as e.g. such as calcium acetate, calcium benzoate, calcium carbonate, calcium chloride, calcium citrate, calcium sorbate, calcium sulfate, potassium ace- tate, potassium benzoate, potassium carbonate, potassium chloride, potassium citrate, potassi- um sorbate, potassium sulfate, sodium acetate, sodium benzoate, sodium carbonate, sodium chloride, sodium citrate, sodium sulfate, zinc acetate, zinc benzoate, zinc carbonate, zinc chlo- ride, zinc citrate, zinc sorbate, zinc sulfate), starch or a sugar or sugar derivative (such as e.g. sucrose, dextrin, glucose, lactose, sorbitol).

In an embodiment, the solid composition is in granulated form. The granule may have a matrix structure where the components are mixed homogeneously. However, the granule typically comprises a core particle and one or more coatings, which typically are salt and/or wax coat- ings. Examples of waxes are polyethylene glycols; polypropylenes; Carnauba wax; Candelilla wax; bees wax; hydrogenated plant oil or animal tallow such as hydrogenated ox tallow, hydro- genated palm oil, hydrogenated cotton seeds and/or hydrogenated soy bean oil; fatty acid alco- hols; mono-glycerides and/or di-glycerides, such as glyceryl stearate, wherein stearate is a mix- ture of stearic and palmitic acid; micro-crystalline wax; paraffin’s; and fatty acids, such as hy- drogenated linear long chained fatty acids and derivatives thereof. A preferred wax is palm oil or hydrogenated palm oil. The core particle can either be a homogeneous blend of galactanase of the invention optionally combined with one or more additional enzymes and optionally to- gether with one or more salts or an inert particle with the galactanase of the invention optionally combined with one or more additional enzymes applied onto it. In an embodiment, the material of the core particles are selected from the group consisting of inorganic salts (such as calcium acetate, calcium benzoate, calcium carbonate, calcium chlo- ride, calcium citrate, calcium sorbate, calcium sulfate, potassium acetate, potassium benzoate, potassium carbonate, potassium chloride, potassium citrate, potassium sorbate, potassium sul- fate, sodium acetate, sodium benzoate, sodium carbonate, sodium chloride, sodium citrate, so- dium sulfate, zinc acetate, zinc benzoate, zinc carbonate, zinc chloride, zinc citrate, zinc sorb- ate, zinc sulfate), starch or a sugar or sugar derivative (such as e.g. sucrose, dextrin, glucose, lactose, sorbitol), sugar or sugar derivative (such as e.g. sucrose, dextrin, glucose, lactose, sor- bitol), small organic molecules, starch, flour, cellulose and minerals and clay minerals (also known as hydrous aluminium phyllosilicates). In a preferred embodiment, the core comprises a clay mineral such as kaolinite or kaolin.

The salt coating is typically at least 1 pm thick and can either be one particular salt or a mixture of salts, such as Na₂S0₄, K₂SO₄, MgS0₄ and/or sodium citrate. Other examples are those de- scribed in e.g. WO 2008/017659, WO 2006/034710, WO 1997/05245, WO 1998/54980, WO 1998/55599, WO 2000/70034 or polymer coating such as described in WO 2001/00042.

In another embodiment, the composition is a solid composition comprising the galactanase of the invention and one or more formulating agents selected from the list consisting of sodium chloride, sodium benzoate, potassium sorbate, sodium sulfate, potassium sulfate, magnesium sulfate, sodium thiosulfate, calcium carbonate, sodium citrate, dextrin, glucose, sucrose, sorbi- tol, lactose, starch and cellulose. In a preferred embodiment, the formulating agent is selected from one or more of the following compounds: sodium sulfate, dextrin, cellulose, sodium thiosul fate and calcium carbonate. In a preferred embodiment, the solid composition is in granulated form. In an embodiment, the solid composition is in granulated form and comprises a core parti- cle, an enzyme layer comprising the galactanase of the invention and a salt coating.

In a further embodiment, the formulating agent is selected from one or more of the following compounds: glycerol, ethylene glycol, 1 , 2-propylene glycol or 1 , 3-propylene glycol, sodium chloride, sodium benzoate, potassium sorbate, sodium sulfate, potassium sulfate, magnesium sulfate, sodium thiosulfate, calcium carbonate, sodium citrate, dextrin, glucose, sucrose, sorbi- tol, lactose, starch, kaolin and cellulose. In a preferred embodiment, the formulating agent is selected from one or more of the following compounds: 1 , 2-propylene glycol, 1 , 3-propylene glycol, sodium sulfate, dextrin, cellulose, sodium thiosulfate, kaolin and calcium carbonate.

Animal Feed and Animal Feed Additives

The present invention also relates to animal feed compositions and animal feed additives corn- prising one or more of the galactanases of the invention. In a further embodiment, the formulat- ing agent comprises one or more of the following compounds: glycerol, ethylene glycol, 1 , 2- propylene glycol or 1 , 3-propylene glycol, sodium chloride, sodium benzoate, potassium sorb- ate, sodium sulfate, potassium sulfate, magnesium sulfate, sodium thiosulfate, calcium car- bonate, sodium citrate, dextrin, glucose, sucrose, sorbitol, lactose, starch, kaolin and cellulose.

Animal feed compositions or diets have a relatively high content of protein. Poultry and pig diets can be characterised as indicated in Table B of WO 01/58275, columns 2-3. Fish diets can be characterised as indicated in column 4 of this Table B. Furthermore such fish diets usually have a crude fat content of 200-310 g/kg.

An animal feed composition according to the invention has a crude protein content of 50-800 g/kg, and can furthermore comprise at least one galactanase of the invention as claimed herein.

Furthermore, or in the alternative (to the crude protein content indicated above), the animal feed composition of the invention has a content of metabolisable energy of 10-30 MJ/kg; and/or a content of calcium of 0.1-200 g/kg; and/or a content of available phosphorus of 0.1-200 g/kg; and/or a content of methionine of 0.1-100 g/kg; and/or a content of methionine plus cysteine of 0.1-150 g/kg; and/or a content of lysine of 0.5-50 g/kg.

In particular embodiments, the content of metabolisable energy, crude protein, calcium, phos- phorus, methionine, methionine plus cysteine, and/or lysine is within any one of ranges 2, 3, 4 or 5 in Table B of WO 01/58275 (R. 2-5).

Crude protein is calculated as nitrogen (N) multiplied by a factor 6.25, i.e. Crude protein (g/kg)= N (g/kg) x 6.25. The nitrogen content is determined by the Kjeldahl method (A.O.A.C., 1984, Official Methods of Analysis 14th ed., Association of Official Analytical Chemists, Washington DC).

Metabolisable energy can be calculated on the basis of the NRC publication Nutrient require- ments in swine, ninth revised edition 1988, subcommittee on swine nutrition, committee on ani- mal nutrition, board of agriculture, national research council. National Academy Press, Washing- ton, D.C., pp. 2-6, and the European Table of Energy Values for Poultry Feed-stuffs, Spelder- holt centre for poultry research and extension, 7361 DA Beekbergen, The Netherlands. Grafisch bedrijf Ponsen & looijen bv, Wageningen. ISBN 90-71463-12-5.

The dietary content of calcium, available phosphorus and amino acids in complete animal diets is calculated on the basis of feed tables such as Veevoedertabel 1997, gegevens over chem- ische samenstelling, verteerbaarheid en voederwaarde van voedermiddelen, Central Veevoed- erbureau, Runderweg 6, 8219 pk Lelystad. ISBN 90-72839-13-7.

In a particular embodiment, the animal feed composition of the invention contains at least one vegetable protein as defined above.

The animal feed composition of the invention may also contain animal protein, such as Meat and Bone Meal, Feather meal, and/or Fish Meal, typically in an amount of 0-25%. The animal feed composition of the invention may also comprise Dried Distillers Grains with Solubles (DDGS), typically in amounts of 0-30%.

The animal feed composition of the invention may also contain insect protein, such as protein from mealworm, housefly or black soldier fly larvae, typically in meal form. Insect meal may re- place fishmeal entirely or in part, and thus may constitute 0-10% of the total feed.

In still further particular embodiments, the animal feed composition of the invention contains 0- 80% maize; and/or 0-80% sorghum; and/or 0-70% wheat; and/or 0-70% Barley; and/or 0-30% oats; and/or 0-40% soybean meal; and/or 0-25% fish meal; and/or 0-25% meat and bone meal; and/or 0-20% whey.

The animal feed may comprise vegetable proteins. In particular embodiments, the protein con- tent of the vegetable proteins is at least 10, 20, 30, 40, 50, 60, 70, 80, or 90% (w/w). Vegetable proteins may be derived from vegetable protein sources, such as legumes and cereals, for ex- ample, materials from plants of the families Fabaceae ( Leguminosae ), Brassicaceae, Amaran- thaceae, and Poaceae, such as soybean meal, lupin meal, rapeseed meal, and combinations thereof.

In a particular embodiment, the vegetable protein source is material from one or more plants of the family Fabaceae, e.g., soybean, lupine, pea, or bean. In another particular embodiment, the vegetable protein source is material from one or more plants of the family Amaranthaceae, e.g. beet, sugar beet, spinach or quinoa. Other examples of vegetable protein sources are rape- seed, crambe and cabbage. In another particular embodiment, soybean is a preferred vegeta- ble protein source. Other examples of vegetable protein sources are cereals such as barley, wheat, rye, oat, maize (corn), rice, and sorghum.

Animal diets can e.g. be manufactured as mash feed (non-pelleted) or pelleted feed. Typically, the milled feed-stuffs are mixed and sufficient amounts of essential vitamins and minerals are added according to the specifications for the species in question. Enzymes can be added as solid or liquid enzyme formulations. For example, for mash feed a solid or liquid enzyme formu- lation may be added before or during the ingredient mixing step. For pelleted feed the (liquid or solid) galactanase enzyme preparation may also be added before or during the feed ingredient step. Typically a liquid galactanase enzyme preparation comprises a galactanase of the inven- tion optionally with a polyol, such as glycerol, ethylene glycol or propylene glycol, and is added after the pelleting step, such as by spraying the liquid formulation onto the pellets. The enzyme may also be incorporated in a feed additive or premix.

Alternatively, the galactanase can be prepared by freezing a mixture of liquid enzyme solution with a bulking agent such as ground soybean meal, and then lyophilizing the mixture.

In an embodiment, the animal feed or animal feed additive comprises one or more additional enzymes. In an embodiment, the animal feed comprises one or more microbes. In an embodi- merit, the animal feed comprises one or more vitamins. In an embodiment, the animal feed comprises one or more minerals. In an embodiment, the animal feed comprises one or more amino acids. In an embodiment, the animal feed comprises one or more other feed ingredients.

In another embodiment, the animal feed or animal feed additive comprises the polypeptide of the invention, one or more formulating agents and one or more additional enzymes. In an em- bodiment, the animal feed or animal feed additive comprises the polypeptide of the invention, one or more formulating agents and one or more microbes. In an embodiment, the animal feed comprises the polypeptide of the invention, one or more formulating agents and one or more vitamins. In an embodiment, the animal feed or animal feed additive comprises one or more minerals. In an embodiment, the animal feed or animal feed additive comprises the polypeptide of the invention, one or more formulating agents and one or more amino acids. In an embodi- ment, the animal feed or animal feed additive comprises the polypeptide of the invention, one or more formulating agents and one or more other feed ingredients.

In a further embodiment, the animal feed or animal feed additive comprises the polypeptide of the invention, one or more formulating agents and one or more components selected from the list consisting of: one or more additional enzymes; one or more microbes; one or more vitamins; one or more minerals; one or more amino acids; and one or more other feed ingredients.

The final enzyme concentration in the diet is within the range of 0.01-200 mg enzyme protein per kg diet, preferably between 0.05-100 mg/kg diet, more preferably 0.1-50 mg, even more preferably 0.2-20 mg enzyme protein per kg animal diet, for each enzyme.

It is at present contemplated that the galactanase is administered in one or more of the follow- ing amounts (dosage ranges): 0.01-200; 0.05-100; 0.1-50; 0.2-20; 0.1-1 ; 0.2-2; 0.5-5; or 1-10 wherein all these ranges are mg galactanase protein per kg feed (ppm).

For determining mg galactanase protein per kg feed, the galactanase is purified from the feed composition, and the specific activity of the purified galactanase is determined using a relevant assay (see under galactanase activity).

In a particular embodiment, the animal feed additive of the invention is intended for being in- cluded (or prescribed as having to be included) in animal diets or feed at levels of 0.01 to 10.0%; more particularly 0.05 to 5.0%; or 0.2 to 1.0% (% meaning g additive per 100 g feed). This is so in particular for premixes.

The same principles apply for determining mg galactanase protein in feed additives. Of course, if a sample is available of the galactanase used for preparing the feed additive or the feed, the specific activity is determined from this sample (no need to purify the galactanase from the feed composition or the additive). Additional Enzymes

In another embodiment, the compositions described herein optionally include one or more addi- tional enzymes. Enzymes can be classified on the basis of the handbook Enzyme Nomencla- ture from NC-IUBMB, 1992), see also the ENZYME site at the internet: http://www.expasy.ch/enzyme/. ENZYME is a repository of information relative to the nomen- clature of enzymes. It is primarily based on the recommendations of the Nomenclature Commit- tee of the International Union of Biochemistry and Molecular Biology (IUB-MB), Academic Press, Inc., 1992, and it describes each type of characterized enzyme for which an EC (Enzyme Commission) number has been provided (Bairoch A. The ENZYME database, 2000, Nucleic Acids Res 28:304-305). This IUB-MB Enzyme nomenclature is based on their substrate speci- ficity and occasionally on their molecular mechanism; such a classification does not reflect the structural features of these enzymes.

Another classification of certain glycoside hydrolase enzymes, such as endoglucanase, alpha- galactosidase, galactanase, mannanase, dextranase, lysozyme and galactosidase is described in Henrissat et al,“The carbohydrate-active enzymes database (CAZy) in 2013”, Nucl. Acids Res. (1 January 2014) 42 (D1 ): D490-D495; see also www.cazy.org.

Thus the composition of the invention may also comprise at least one other enzyme selected from the group comprising of phytase (EC 3.1.3.8 or 3.1.3.26); xylanase (EC 3.2.1.8); galac- tanase (EC 3.2.1.89); alpha-galactosidase (EC 3.2.1.22); protease (EC 3.4); phospholipase A1 (EC 3.1.1.32); phospholipase A2 (EC 3.1.1.4); lysophospholipase (EC 3.1.1.5); phospholipase C (3.1.4.3); phospholipase D (EC 3.1.4.4); amylase such as, for example, alpha-amylase (EC 3.2.1.1 ); arabinofuranosidase (EC 3.2.1.55); beta-xylosidase (EC 3.2.1.37); acetyl xylan ester- ase (EC 3.1.1.72); feruloyl esterase (EC 3.1.1.73); cellulase (EC 3.2.1.4); cellobiohydrolases (EC 3.2.1.91 ); beta-glucosidase (EC 3.2.1.21 ); pullulanase (EC 3.2.1.41 ), alpha-mannosidase (EC 3.2.1.24), mannanase (EC 3.2.1.25) and beta-glucanase (EC 3.2.1.4 or EC 3.2.1.6), or any mixture thereof.

In a particular embodiment, the composition of the invention comprises a phytase (EC 3.1.3.8 or 3.1.3.26). Examples of commercially available phytases include Bio-Feed™ Phytase (Novo- zymes), Ronozyme® P, Ronozyme® NP and Ronozyme® HiPhos (DSM Nutritional Products), Natuphos® and Naturphos® E (BASF), Finase® and Quantum® Blue (AB Enzymes), OptiPhos® (Huvepharma) Phyzyme® XP (Verenium/DuPont) and Axtra® PHY (DuPont). Other preferred phytases include those described in e.g. WO 98/28408, WO 00/43503, and WO 03/066847.

In a particular embodiment, the composition of the invention comprises a xylanase (EC 3.2.1.8). Examples of commercially available xylanases include Ronozyme® WX and Ronozyme® G2 (DSM Nutritional Products), Econase® XT and Barley (AB Vista), Xylathin® (Verenium), Hos- tazym® X (Huvepharma) and Axtra® XB (xylanase /beta-glucanase, DuPont). In a particular embodiment, the composition of the invention comprises a protease (EC 3.4). Examples of commercially available proteases include Ronozyme® ProAct (DSM Nutritional Products).

Microbes

In an embodiment, the composition, animal feed additive or animal feed further comprises one or more additional microbes. In a particular embodiment, the composition, animal feed additive or animal feed further comprises a bacterium from one or more of the following genera: Lacto bacillus, Lactococcus, Streptococcus, Bacillus, Pediococcus, Enterococcus, Leuconostoc, Car- nobacterium, Propionibacterium, Bifidobacterium, Clostridium and Megasphaera or any combi- nation thereof.

In a preferred embodiment, composition, animal feed additive or animal feed further comprises a bacterium from one or more of the following strains: Bacillus subtilis, Bacillus licheniformis, Bacillus amyloliquefaciens, Bacillus cereus, Bacillus pumilus, Bacillus polymyxa, Bacillus mega- terium, Bacillus coagulans, Bacillus circulans, Enterococcus faecium, Enterococcus spp, and Pediococcus spp, Lactobacillus spp, Bifidobacterium spp, Lactobacillus acidophilus, Pedio- cocsus acidilactici, Lactococcus lactis, Bifidobacterium bifidum, Propionibacterium thoenii, Lac tobacillus farciminus, lactobacillus rhamnosus, Clostridium butyricum, Bifidobacterium animalis ssp. animalis, Lactobacillus reuteri, Lactobacillus salivarius ssp. salivarius, Megasphaera els- denii, Propionibacteria sp.

In a more preferred embodiment, composition, animal feed additive or animal feed further corn- prises a bacterium from one or more of the following strains of Bacillus subtilis: 3A-P4 (PTA- 6506); 15A-P4 (PTA-6507); 22C-P1 (PTA-6508); 2084 (NRRL B-500130); LSSA01 (NRRL-B- 50104); BS27 (NRRL B-501 05); BS 18 (NRRL B-50633); BS 278 (NRRL B-50634); DSM 29870; DSM 29871 ; NRRL B-50136; NRRL B-50605; NRRL B-50606; NRRL B-50622 and PTA- 7547.

In a more preferred embodiment, composition, animal feed additive or animal feed further corn- prises a bacterium from one or more of the following strains of Bacillus pumilus : NRRL B-50016; ATCC 700385; NRRL B-50885; NRRL B-50886

In a more preferred embodiment, composition, animal feed additive or animal feed further corn- prises a bacterium from one or more of the following strains of Bacillus lichenformis : NRRL B 50015; NRRL B-50621 ; NRRL B-50623

In a more preferred embodiment, composition, animal feed additive or animal feed further corn- prises a bacterium from one or more of the following strains of Bacillus amyloliquefaciens: DSM 29869; DSM 29869; NRRL B 50607; PTA-7543; PTA-7549; NRRL B-50349; NRRL B-50606; NRRL B-50013; NRRL B-50151 ; NRRL B-50141 ; NRRL B-50147; NRRL B-50888 The bacterial count of each of the bacterial strains in the composition, animal feed additive or animal feed is between 1x10⁴ and 1x10¹⁴ CFU/kg of dry matter, preferably between 1x10⁶ and 1x10¹² CFU/kg of dry matter, and more preferably between 1x10⁷ and 1x10¹¹ CFU/kg of dry matter. In a more preferred embodiment the bacterial count of each of the bacterial strains in the composition, animal feed additive or animal feed is between 1x10⁸ and 1x10¹⁰ CFU/kg of dry matter.

The bacterial count of each of the bacterial strains in the composition, animal feed additive or animal feed is between 1x10⁵ and 1x10¹⁵ CFU/animal/day, preferably between 1x10⁷ and 1x10¹³ CFU/animal/day, and more preferably between 1x10⁸ and 1x10¹² CFU/animal/day. In a more preferred embodiment the bacterial count of each of the bacterial strains in the composi- tion, animal feed additive or animal feed is between 1x10⁹ and 1x10¹¹ CFU/animal/day.

In another embodiment, the one or more bacterial strains are present in the form of a stable spore.

Premix

In an embodiment, the animal feed may include a premix, comprising e.g. vitamins, minerals, enzymes, amino acids, preservatives, antibiotics, other feed ingredients or any combination thereof which are mixed into the animal feed.

Amino Acids

The composition of the invention may further comprise one or more amino acids. Examples of amino acids which are used in animal feed are lysine, alanine, beta-alanine, threonine, methio- nine and tryptophan.

Vitamins and Minerals

In another embodiment, the animal feed may include one or more vitamins, such as one or more fat-soluble vitamins and/or one or more water-soluble vitamins. In another embodiment, the animal feed may optionally include one or more minerals, such as one or more trace miner- als and/or one or more macro minerals.

Usually fat- and water-soluble vitamins, as well as trace minerals form part of a so-called premix intended for addition to the feed, whereas macro minerals are usually separately added to the feed.

Non-limiting examples of fat-soluble vitamins include vitamin A, vitamin D3, vitamin E, and vita- min K, e.g., vitamin K3.

Non-limiting examples of water-soluble vitamins include vitamin B12, biotin and choline, vitamin B1 , vitamin B2, vitamin B6, niacin, folic acid and panthothenate, e.g., Ca-D-panthothenate. Non-limiting examples of trace minerals include boron, cobalt, chloride, chromium, copper, fluo- ride, iodine, iron, manganese, molybdenum, selenium and zinc.

Non-limiting examples of macro minerals include calcium, magnesium, potassium and sodium.

The nutritional requirements of these components (exemplified with poultry and piglets/pigs) are listed in Table A of WO 01/58275. Nutritional requirement means that these components should be provided in the diet in the concentrations indicated.

In the alternative, the animal feed additive of the invention comprises at least one of the individ- ual components specified in Table A of WO 01/58275. At least one means either of, one or more of, one, or two, or three, or four and so forth up to all thirteen, or up to all fifteen individual components. More specifically, this at least one individual component is included in the additive of the invention in such an amount as to provide an in-feed-concentration within the range indi- cated in column four, or column five, or column six of Table A.

In a still further embodiment, the animal feed additive of the invention comprises at least one of the below vitamins, preferably to provide an in-feed-concentration within the ranges specified in the below Table 1 (for piglet diets, and broiler diets, respectively).

Table 1 : Typical vitamin recommendations

Other feed ingredients

The composition of the invention may further comprise colouring agents, stabilisers, growth im- proving additives and aroma compounds/flavourings, polyunsaturated fatty acids (PUFAs); reac- tive oxygen generating species, anti-microbial peptides and anti-fungal polypeptides.

Examples of colouring agents are carotenoids such as beta-carotene, astaxanthin, and lutein.

Examples of aroma compounds/flavourings are creosol, anethol, deca-, undeca-and/or dodeca- lactones, ionones, irone, gingerol, piperidine, propylidene phatalide, butylidene phatalide, cap- saicin and tannin.

Examples of antimicrobial peptides (AMP’s) are CAP18, Leucocin A, Tritrpticin, Protegrin-1 , Thanatin, Defensin, Lactoferrin, Lactoferricin, and Ovispirin such as Novispirin (Robert Lehrer, 2000), Plectasins, and Statins, including the compounds and polypeptides disclosed in WO 03/044049 and WO 03/048148, as well as variants or fragments of the above that retain antimi- crobial activity.

Examples of antifungal polypeptides (AFP’s) are the Aspergillus giganteus, and Aspergillus ni- ger peptides, as well as variants and fragments thereof which retain antifungal activity, as dis closed in WO 94/01459 and WO 02/090384.

Examples of polyunsaturated fatty acids are C18, C20 and C22 polyunsaturated fatty acids, such as arachidonic acid, docosohexaenoic acid, eicosapentaenoic acid and gamma-linoleic acid.

Examples of reactive oxygen generating species are chemicals such as perborate, persulphate, or percarbonate; and enzymes such as an oxidase, an oxygenase or a syntethase.

The composition of the invention may further comprise at least one amino acid. Examples of amino acids which are used in animal feed are lysine, alanine, beta-alanine, threonine, methio- nine and tryptophan.

Uses

The present invention is also directed to methods (preferably in vitro methods) for using the pol- ypeptides having galactanase activity, or compositions thereof, for e.g. improving the nutritional value of soy-based material. The present invention is also directed to processes for using the polypeptides having galactanase or compositions thereof, such as e.g. those described below. Use in Animal Feed

The present invention is also directed to methods (preferably in vitro methods) for using the ga- lactanases of the invention to modify and/or improve the nutritional value of animal feed such as soy-based materials.

The term animal includes all animals. Examples of animals are non-ruminants, and ruminants. Ruminant animals include, for example, animals such as sheep, goats, and cattle, e.g. beef cat- tie, cows, and young calves. In a particular embodiment, the animal is a non-ruminant animal. Non-ruminant animals include mono-gastric animals, e.g. pigs or swine (including, but not lim ited to, piglets, growing pigs, and sows); poultry such as turkeys, ducks and chicken (including but not limited to broiler chicks, layers); horses (including but not limited to hotbloods, cold- bloods and warm bloods), young calves; and fish (including but not limited to salmon, trout, ti- lapia, catfish and carps; and crustaceans (including but not limited to shrimps and prawns).

In the use according to the invention the galactanases can be used to treat animal feed such as soy-based materials prior to feeding an animal with said feed (or said soy-based materials).

In a particular embodiment, the form in which the galactanase is added to the feed, or animal feed additive, is well-defined. Well-defined means that the galactanase preparation is at least 50% pure as determined by Size-exclusion chromatography (see Example 12 of WO 01/58275). In other particular embodiments the galactanase preparation is at least 60, 70, 80, 85, 88, 90, 92, 94, or at least 95% pure as determined by this method.

A well-defined galactanase preparation is advantageous. For instance, it is much easier to dose correctly to the feed a galactanase that is essentially free from interfering or contaminating other galactanases. The term dose correctly refers in particular to the objective of obtaining consistent and constant results, and the capability of optimizing dosage based upon the desired effect.

For the use in animal feed, however, the galactanase need not be that pure; it may e.g. include other enzymes, in which case it could be termed a galactanase preparation.

PREFERRED EMBODIMENTS

Preferred embodiments of the invention are described in the set of items below.

1. A method for modifying and/or improving the nutritional value of a soy-based material, the method comprising the steps of

i) providing a soy-based material and

ii) treating the soy-based material with a polypeptide having galactanase activity selected from the group consisting of: (a) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 3;

(b) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 7;

(c) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 1 1 ;

(d) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 15;

(e) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 19;

(f) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 23;

(g) a polypeptide encoded by a polynucleotide that hybridizes under high stringency conditions, or very high stringency conditions with

(vi) the mature polypeptide coding sequence of SEQ ID NO: 21 or the cDNA sequence thereof, (vii) the full-length complementary strand of (i), (ii), (iii), (iv), (v) or (vi);

(h) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 1 ;

(i) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 5;

(j) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 9;

(k) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 13;

(l) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 17;

(m) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 21 ;

(n) a variant of SEQ ID NO: 3 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(o) a variant of SEQ ID NO: 7 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(p) a variant of SEQ ID NO: 11 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(q) a variant of SEQ ID NO: 15 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(r) a variant of SEQ ID NO: 19 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(s) a variant of SEQ ID NO: 23 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(t) a polypeptide comprising the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (I), (m), (n), (o), (p), (q), (r) or (s) and a N-terminal and/or C-terminal His-tag and/or HQ-tag; and

(u) a fragment of the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (I), (m), (n), (o), (p), (q), (r), (s) or (t) having galactanase activity and having at least 90% of the length of the mature polypeptide,

wherein the polypeptide optionally has galactanase activity which is at least 60%, such as at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95% or at least 100% of the galactanase activity of the polypeptide of SEQ ID NO: 3.

2. A method for modifying and/or improving the nutritional value of a soy-based material, the method comprising the steps of

i) providing a soy-based material and

(g) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 31 ;

(iv) the mature polypeptide coding sequence of SEQ ID NO: 13 or the cDNA sequence thereof, (v) the mature polypeptide coding sequence of SEQ ID NO: 17 or the cDNA sequence thereof,

(vii) the mature polypeptide coding sequence of SEQ ID NO: 29 or the cDNA sequence thereof,

(viii)the full-length complementary strand of (i), (ii), (iii), (iv), (v), (vi) or (vii);

(i) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 1 ;

(j) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 5;

(k) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 9;

(l) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 13;

(m) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 17;

(n) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 21 ; (o) a variant of SEQ ID NO: 3 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(p) a variant of SEQ ID NO: 7 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(v) a polypeptide comprising the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (I), (m), (n), (o), (p), (q), (r), (s), (t), or (u) and a N-terminal and/or C-terminal His-tag and/or HQ-tag; and (w) a fragment of the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (I), (m), (n),

(o), (p), (q), (r), (s), (t), (u) or (v) having galactanase activity and having at least 90% of the length of the mature polypeptide,

3. The method according to item 1 or 2, wherein the polypeptide is a GH 53 polypeptide having galactanase activity.

4. The method according to any of the previous items, wherein the polypeptide has beta galactanase activity.

5. The method according to any of the previous items, wherein the soy-based material is selected from the group consisting of dehulled full-fat flakes, full-fat soybean meal, defatted flakes, white flakes, full-fat flakes and soybean meal.

6. The method according to any of the previous items, wherein the soy-based material is in an aqueous solution.

7. The method according to any of the previous items, wherein the soy-based material has a dry-matter content of from 40-60% (w/w) such as from 40%-45% (w/w), for example from 45%- 50 (w/w) such as from 50%-55% (w/w), for example from 55%-60 (w/w), or any combination of these intervals.

8. The method according to any of items 1 to 6, wherein the soy-based material has a dry- matter content of from 20-40% (w/w) such as from 20-25% (w/w), for example from 25-30% (w/w), such as from 30-35% (w/w), for example from 35-40% (w/w) or any combination of these intervals.

9. The method according to any of the previous items, wherein step ii) comprises galactanase treatment of the soy-based material for 24 hours to 5 days such as from 2 days to 4 days.

10. The method according to any of items 1 to 8, wherein step ii) comprises galactanase treatment of the soy-based material for 6 hours to 24 hours such as from 10 hours to 20 hours.

1 1. The method according to any of items 1 to 8, wherein step ii) comprises galactanase treatment of the soy-based material for 30 minutes to 6 hours such as from 1 hour to 5 hours. 12. The method according to any of the previous items, wherein step ii) comprises use of from 0.001 mg galactanase /dry-matter to 6 mg galactanase /dry-matter such as from 0.001 mg galactanase /dry-matter to 0.005 mg galactanase /dry-matter, for example from 0.005 mg galactanase /dry-matter to 0.01 mg galactanase /dry-matter, such as from 0.01 mg galactanase /dry-matter to 0.02 mg galactanase /dry-matter, for example from 0.02 mg galactanase /dry- matter to 0.05 mg galactanase /dry-matter, such as from 0.05 mg galactanase /dry-matter to 0.1 mg galactanase /dry-matter, for example from 0.1 mg galactanase /dry-matter to 0.5 mg galactanase /dry-matter, such as from 0.5 mg galactanase /dry-matter to 1 mg galactanase /dry- matter, for example from 1 mg galactanase /dry-matter to 2 mg galactanase /dry-matter, such as from 2 mg galactanase /dry-matter to 3 mg galactanase /dry-matter, for example from 3 mg galactanase /dry-matter to 4 mg galactanase /dry-matter, such as from 4 mg galactanase /dry- matter to 5 mg galactanase /dry-matter, for example from 5 mg galactanase /dry-matter to 6 mg galactanase /dry-matter or any combination of these intervals.

13. The method according to any of the previous items wherein step ii) is performed in vitro.

14. The method according to any of the previous items wherein step ii) is performed before one or more alcohol leaching steps.

15. The method according to any of the previous items, wherein an alcohol leaching step is performed after step ii).

16. The method according to any of the previous items, wherein galactanase treated soy-based material has an increased protein content (compared to soy-based material that has not been treated with galactanase).

17. The method according to item 16, wherein the protein content is increased by at least 1% such as at least 2%, such as at least 3%, such as at least 4%, such as at least 5%, such as at least 6%, such as at least 7%, such as at least 8%, such as at least 9%, or such as at least 10%.

18. The method according to any of the previous items, wherein galactanase treated soy-based material has an increased NDF digestibility (compared to soy-based material that has not been treated with galactanase).

19. The method according to item 18, wherein the NDF digestibility is increased by at least 1 % such as at least 2%, such as at least 3%, such as at least 4%, such as at least 5%, such as at least 6%, such as at least 7%, such as at least 8%, such as at least 9%, or such as at least 10%.

20. The method according to any of the previous items, wherein step ii) comprises galactanase treatment of the soy-based material at pH between pH 3 and 6, such as between pH 3 and 4, for example between pH 4 and 5, such as pH 5 and 6, or any combination of these intervals.

21. The method according to any of the previous items, wherein step ii) comprises galactanase treatment of the soy-based material at a temperature between 40°C and 80°C such as between 50°C and 70°C.

22. The method according to any of items 1 to 20, wherein step ii) comprises galactanase treatment of the soy-based material at a temperature between 15°C and 40°C such as between 20°C and 30°C.

23. The method according to any of the previous items, wherein step ii) comprises galactanase treatment and fermentation in a single step.

24. The method according to any of the previous items, wherein step ii) is followed by one or more fermentation steps.

25. The method according to any of the previous items, wherein the method results in production of soybean meal, fermented soybean meal or soy protein concentrate.

26. The method according to any of the previous items, wherein the polypeptide comprises or consists of amino acids 1 to 316 of SEQ ID NO: 2, amino acids 1 to 316 of SEQ ID NO: 3, ami- no acids 1 to 324 of SEQ ID NO: 4, amino acids 1 to 318 of SEQ ID NO: 6, amino acids 1 to 318 of SEQ ID NO: 7, amino acids 1 to 326 of SEQ ID NO: 8, amino acids 1 to 516 of SEQ ID NO: 10, amino acids 1 to 516 of SEQ ID NO: 1 1 , amino acids 1 to 524 of SEQ ID NO: 12, amino ac- ids 1 to 317 of SEQ ID NO: 14, amino acids 1 to 317 of SEQ ID NO: 15, amino acids 1 to 325 of SEQ ID NO: 16, amino acids 1 to 316 of SEQ ID NO: 18, amino acids 1 to 316 of SEQ ID NO: 19, amino acids 1 to 324 of SEQ ID NO: 20, amino acids 1 to 316 of SEQ ID NO: 22, amino ac- ids 1 to 316 of SEQ ID NO: 23 or amino acids 1 to 324 of SEQ ID NO: 24.

27. The method according to any of the previous items, wherein the polypeptide comprises or consists of amino acids 1 to 316 of SEQ ID NO: 2, amino acids 1 to 316 of SEQ ID NO: 3, ami- no acids 1 to 324 of SEQ ID NO: 4, amino acids 1 to 318 of SEQ ID NO: 6, amino acids 1 to 318 of SEQ ID NO: 7, amino acids 1 to 326 of SEQ ID NO: 8, amino acids 1 to 516 of SEQ ID NO: 10, amino acids 1 to 516 of SEQ ID NO: 1 1 , amino acids 1 to 524 of SEQ ID NO: 12, amino ac- ids 1 to 317 of SEQ ID NO: 14, amino acids 1 to 317 of SEQ ID NO: 15, amino acids 1 to 325 of SEQ ID NO: 16, amino acids 1 to 316 of SEQ ID NO: 18, amino acids 1 to 316 of SEQ ID NO: 19, amino acids 1 to 324 of SEQ ID NO: 20, amino acids 1 to 316 of SEQ ID NO: 22, amino ac- ids 1 to 316 of SEQ ID NO: 23 or amino acids 1 to 324 of SEQ ID NO: 24, amino acids 1 to 307 of SEQ ID NO: 30, amino acids 1 to 307 of SEQ ID NO: 31 or amino acids 1 to 315 of SEQ ID NO: 32.

28. The method according to any of the previous items, wherein the polypeptide comprises the motif GV[T/M]PD[W/M]VQ[I/V]GNE (SEQ ID NO: 26) and/or the motif WADP[A/G]xQxKPxAW (SEQ ID NO: 27).

29. An animal feed or animal feed additive comprising the galactanase treated soy-based material obtained in the method according to any of items 1 to 28.

30. An isolated polypeptide having galactanase activity, selected from the group consisting of:

(a) a polypeptide having at least 82%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 3;

(b) a polypeptide having at least 83%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 7;

(c) a polypeptide having at least 86%, e.g., at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 1 1 ;

(d) a polypeptide having at least 99.3%, e.g., at least 99.6% or 100% sequence identity to the polypeptide of SEQ ID NO: 15;

(e) a polypeptide having at least 99.3%, e.g., at least 99.6% or 100% sequence identity to the polypeptide of SEQ ID NO: 19;

(f) a polypeptide having at least 83%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 23;

(g) a polypeptide encoded by a polynucleotide that hybridizes under high stringency conditions, or very high stringency conditions with (i) the mature polypeptide coding sequence of SEQ ID NO: 1 or the cDNA sequence thereof,

(vii) the full-length complementary strand of (i), (ii), (iii), (iv), (v) or (vi);

(h) a polypeptide encoded by a polynucleotide having at least 82%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 1 ;

(i) a polypeptide encoded by a polynucleotide having at least 83%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 5;

(j) a polypeptide encoded by a polynucleotide having at least 86%, e.g., at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 9;

(k) a polypeptide encoded by a polynucleotide having at least 99.3%, e.g., at least 99.6% or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 13;

(l) a polypeptide encoded by a polynucleotide having at least 99.3%, e.g., at least 99.6% or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 17;

(m) a polypeptide encoded by a polynucleotide having at least 83%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 21 ; (n) a variant of SEQ ID NO: 3 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(p) a variant of SEQ ID NO: 1 1 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(q) a variant of SEQ ID NO: 15 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 or 2 positions;

(r) a variant of SEQ ID NO: 19 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 or 2 positions;

(t) a polypeptide comprising the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (I), (m), (n), (o), (p), (q), (r) or (s) and a N-terminal and/or C-terminal His-tag and/or HQ- tag; and

(u) a fragment of the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (I), (m), (n), (o), (p), (q), (r), (s) or (t) having galactanase activity and having at least 90% of the length of the mature polypeptide, wherein the polypeptide optionally has galactanase activity which is at least 60%, such as at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95% or at least 100% of the galactanase activity of the polypeptide of SEQ ID NO: 3.

31. An isolated polypeptide having galactanase activity, selected from the group consisting of:

(g) a polypeptide having at least 95%, e.g. at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5% or 100% sequence identity to the polypeptide of SEQ ID NO: 31 ;

(viii) the full-length complementary strand of (i), (ii), (iii), (iv), (v), (vi) or (vii);

(i) a polypeptide encoded by a polynucleotide having at least 82%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 1 ;

(j) a polypeptide encoded by a polynucleotide having at least 83%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 5;

(k) a polypeptide encoded by a polynucleotide having at least 86%, e.g., at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 9;

(l) a polypeptide encoded by a polynucleotide having at least 99.3%, e.g., at least 99.6% or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 13;

(m) a polypeptide encoded by a polynucleotide having at least 99.3%, e.g., at least 99.6% or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 17;

(n) a polypeptide encoded by a polynucleotide having at least 83%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 21 ;

(o) a polypeptide encoded by a polynucleotide having at least 95%, e.g. at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5% or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 29;

(p) a variant of SEQ ID NO: 3 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(q) a variant of SEQ ID NO: 7 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(r) a variant of SEQ ID NO: 1 1 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(s) a variant of SEQ ID NO: 15 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 or 2 positions;

(t) a variant of SEQ ID NO: 19 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 or 2 positions;

(u) a variant of SEQ ID NO: 23 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(v) a variant of SEQ ID NO: 31 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13 ,14, 15 or 16 positions;

(w) a polypeptide comprising the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (I), (m), (n), (o), (p), (q), (r), (s), (t), (u) or (v) and a N-terminal and/or C-terminal His- tag and/or HQ-tag; and (x) a fragment of the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (I), (m), (n), (o),

(p), (q), (r), (s), (t), (u), (v) or (x) having galactanase activity and having at least 90% of the length of the mature polypeptide,

32. The polypeptide according to item 30, wherein the polypeptide comprises or consists of amino acids 1 to 316 of SEQ ID NO: 2, amino acids 1 to 316 of SEQ ID NO: 3, amino acids 1 to 324 of SEQ ID NO: 4, amino acids 1 to 318 of SEQ ID NO: 6, amino acids 1 to 318 of SEQ ID

NO: 7, amino acids 1 to 326 of SEQ ID NO: 8, amino acids 1 to 516 of SEQ ID NO: 10, amino acids 1 to 516 of SEQ ID NO: 1 1 , amino acids 1 to 524 of SEQ ID NO: 12, amino acids 1 to 317 of SEQ ID NO: 14, amino acids 1 to 317 of SEQ ID NO: 15, amino acids 1 to 325 of SEQ ID NO: 16, amino acids 1 to 316 of SEQ ID NO: 18, amino acids 1 to 316 of SEQ ID NO: 19, amino acids 1 to 324 of SEQ ID NO: 20, amino acids 1 to 316 of SEQ ID NO: 22, amino acids 1 to 316 of SEQ ID NO: 23 or amino acids 1 to 324 of SEQ ID NO: 24.

33. The polypeptide according to item 31 , wherein the polypeptide comprises or consists of amino acids 1 to 316 of SEQ ID NO: 2, amino acids 1 to 316 of SEQ ID NO: 3, amino acids 1 to 324 of SEQ ID NO: 4, amino acids 1 to 318 of SEQ ID NO: 6, amino acids 1 to 318 of SEQ ID

NO: 7, amino acids 1 to 326 of SEQ ID NO: 8, amino acids 1 to 516 of SEQ ID NO: 10, amino acids 1 to 516 of SEQ ID NO: 11 , amino acids 1 to 524 of SEQ ID NO: 12, amino acids 1 to 317 of SEQ ID NO: 14, amino acids 1 to 317 of SEQ ID NO: 15, amino acids 1 to 325 of SEQ ID NO: 16, amino acids 1 to 316 of SEQ ID NO: 18, amino acids 1 to 316 of SEQ ID NO: 19, amino acids 1 to 324 of SEQ ID NO: 20, amino acids 1 to 316 of SEQ ID NO: 22, amino acids 1 to 316 of SEQ ID NO: 23, amino acids 1 to 324 of SEQ ID NO: 24, amino acids 1 to 307 of SEQ ID NO: 30, amino acids 1 to 307 of SEQ ID NO: 31 or amino acids 1 to 315 of SEQ ID NO: 32.

34. The polypeptide according to any of items 30 to 33, wherein the polypeptide comprises the motif GV[T/M]PD[W/M]VQ[IA/]GNE (SEQ ID NO: 26) and/or the motif WADP[A/G]xQxKPxAW (SEQ ID NO: 27).

35. A polynucleotide encoding the polypeptide of any of items 30 to 34.

36. A nucleic acid construct or expression vector comprising the polynucleotide of item 35 oper- ably linked to one or more control sequences that direct the production of the polypeptide in an expression host. 37. A recombinant host cell comprising the polynucleotide of item 35 operably linked to one or more control sequences that direct the production of the polypeptide.

38. A method of producing the polypeptide of any of items 30 to 34, comprising:

(a) cultivating a cell, which in its wild-type form produces the polypeptide, under conditions conductive for production of the polypeptide; and

(b) recovering the polypeptide.

39. A method of producing the polypeptide of any of items 30 to 34, comprising:

(a) cultivating the recombinant host cell of item 37 under conditions conducive for production of the polypeptide; and

(b) recovering the polypeptide.

40. A transgenic plant, plant part or plant cell transformed with a polynucleotide encoding the polypeptide of any of items 30 to 34.

41. A whole broth formulation or cell culture composition comprising a polypeptide of any of items 30 to 34.

42. A composition comprising the polypeptide of any of items 30 to 34.

43. The composition of item 42 further comprising one or more formulating agents.

44. The composition of item 43, wherein the one or more formulating agent is selected from the group consisting of glycerol, ethylene glycol, 1 , 2-propylene glycol or 1 , 3-propylene glycol, so- dium chloride, sodium benzoate, potassium sorbate, sodium sulfate, potassium sulfate, magne- sium sulfate, sodium thiosulfate, calcium carbonate, sodium citrate, dextrin, glucose, sucrose, sorbitol, lactose, starch and cellulose or any combination thereof.

45. The composition of any of items 42 to 44 which is in granulate form.

46. The composition of item 45 wherein the granulate is coated.

47. The composition of item 46 wherein the coating comprises a salt and/or wax.

48. The composition of any of items 42 to 47 further comprising one or more additional en- zymes. 49. The composition of item 48 wherein the one or more additional enzymes is selected from the group consisting of phytase, galactanase, alpha-galactosidase, protease, phospholipase A1 , phospholipase A2, lysophospholipase, phospholipase C, phospholipase D, amylase, lysozyme, beta-galactosidase, beta-xylosidase, acetyl xylan esterase, feruloyl esterase, cellulase, cellobi- ohydrolases, beta-glucosidase, pullulanase, and beta-glucanase or any combination thereof.

50. Use of an isolated polypeptide having galactanase activity for improving the nutritional value of soy-based material, wherein the polypeptide is selected from the group consisting of:

(a) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 3;

(ii) the mature polypeptide coding sequence of SEQ ID NO: 5 or the cDNA sequence thereof, (iii) the mature polypeptide coding sequence of SEQ ID NO: 9 or the cDNA sequence thereof,

(m) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 21 ;

(s) a variant of SEQ ID NO: 23 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions; (t) a polypeptide comprising the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), Q), (k), (I), (m), (n), (o), (p), (q), (r) or (s) and a N-terminal and/or C-terminal His-tag and/or HQ- tag; and

(u) a fragment of the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (I), (m), (n), (o),

(р), (q), (r), (s) or (t) having galactanase activity and having at least 90% of the length of the mature polypeptide,

51. Use of an isolated polypeptide having galactanase activity for improving the nutritional value of soy-based material, wherein the polypeptide is selected from the group consisting of:

(с) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 1 1 ;

(n) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 21 ;

(o) a variant of SEQ ID NO: 3 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(q) a variant of SEQ ID NO: 1 1 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(v) a polypeptide comprising the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (I), (m), (n), (o), (p), (q), (r), (s), (t) or (u) and a N-terminal and/or C-terminal His-tag and/or HQ-tag; and

(w) a fragment of the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (I), (m), (n), (o), (p), (q), (r), (s), (t), (u) or (v) having galactanase activity and having at least 90% of the length of the mature polypeptide,

52. The use according to item 50, wherein the polypeptide comprises or consists of amino acids 1 to 316 of SEQ ID NO: 2, amino acids 1 to 316 of SEQ ID NO: 3, amino acids 1 to 324 of SEQ ID NO: 4, amino acids 1 to 318 of SEQ ID NO: 6, amino acids 1 to 318 of SEQ ID NO: 7, amino acids 1 to 326 of SEQ ID NO: 8, amino acids 1 to 516 of SEQ ID NO: 10, amino acids 1 to 516 of SEQ ID NO: 1 1 , amino acids 1 to 524 of SEQ ID NO: 12, amino acids 1 to 317 of SEQ ID NO: 14, amino acids 1 to 317 of SEQ ID NO: 15, amino acids 1 to 325 of SEQ ID NO: 16, amino acids 1 to 316 of SEQ ID NO: 18, amino acids 1 to 316 of SEQ ID NO: 19, amino acids 1 to 324 of SEQ ID NO: 20, amino acids 1 to 316 of SEQ ID NO: 22, amino acids 1 to 316 of SEQ ID NO: 23 or amino acids 1 to 324 of SEQ ID NO: 24.

53. The use according to item 51 , wherein the polypeptide comprises or consists of amino acids 1 to 316 of SEQ ID NO: 2, amino acids 1 to 316 of SEQ ID NO: 3, amino acids 1 to 324 of SEQ ID NO: 4, amino acids 1 to 318 of SEQ ID NO: 6, amino acids 1 to 318 of SEQ ID NO: 7, amino acids 1 to 326 of SEQ ID NO: 8, amino acids 1 to 516 of SEQ ID NO: 10, amino acids 1 to 516 of SEQ ID NO: 1 1 , amino acids 1 to 524 of SEQ ID NO: 12, amino acids 1 to 317 of SEQ ID NO: 14, amino acids 1 to 317 of SEQ ID NO: 15, amino acids 1 to 325 of SEQ ID NO: 16, amino acids 1 to 316 of SEQ ID NO: 18, amino acids 1 to 316 of SEQ ID NO: 19, amino acids 1 to 324 of SEQ ID NO: 20, amino acids 1 to 316 of SEQ ID NO: 22, amino acids 1 to 316 of SEQ ID NO: 23, amino acids 1 to 324 of SEQ ID NO: 24, amino acids 1 to 307 of SEQ ID NO: 30, amino acids 1 to 307 of SEQ ID NO: 31 or amino acids 1 to 315 of SEQ ID NO: 32.

54. The use according to any of item 50 to 53, wherein the polypeptide comprises the motif GV[T/M]PD[W/M]VQ[I/V]GNE (SEQ ID NO: 26) and/or the motif WADP[A/G]xQxKPxAW (SEQ ID NO: 27).

EXAMPLES

Strains

The galactanases were derived from bacterial strains isolated from environmental samples by standard microbiological isolation techniques. Strains were identified and taxonomy was as- signed based on DNA sequencing of the 16S ribosomal genes (Table 1 ).

Table 1 : Isolation of bacterial strains

Chromosomal DNA isolated from pure cultures of the individual strains with the DNeasy Blood & Tissue Kit from Qiagen (Hilden, Germany) was subjected to full genome sequencing using lllu- mina technology. Genome sequencing, the subsequent assembly of reads and the gene dis- covery (i.e. annotation of gene functions) is known to the person skilled in the art and the ser- vice can be purchased commercially. The bacterial genome sequences were analyzed for ga- lactanases from the CAZY database family GH53 (Lombard et al. The Carbohydrate-active en- zymes database CAZy. Nucleic Acids Res 2013, 42:D490-D495.) This analysis identified ten gene encoding putative galactanases with the nucleotide sequences given in SEQ ID NO: 1 , 5, 9, 13, 17 and 21.

Example 1 : Cloning of GH53 galactanases from Cohnella sp-60555 (SEQ ID NO: 4), Cohnella xylanilytica (SEQ ID NO: 8), Paenibacillus sp-18179 (SEQ ID NO: 12), Paenibacil- lus peoriae (SEQ ID NO: 16), Paenibacillus xylanexedens (SEQ ID NO: 20) and Cohnella laeviribosi (SEQ ID NO: 24)

The genes encoding the galactanases were amplified by PCR and fused with regulatory ele- ments, affinity purification tag and homology regions for recombination into the B. subtilis ge- nome. The linear integration construct was a SOE-PCR fusion product (Horton, R.M., Hunt, H.D., Ho, S.N., Pullen, J.K. and Pease, L.R. (1989) Engineering hybrid genes without the use of restriction enzymes, gene splicing by overlap extension Gene 77: 61-68) made by fusion of the gene between two Bacillus subtilis chromosomal regions along with strong promoters and a chloramphenicol resistance marker. The SOE PCR method is also described in patent applica- tion WO 2003095658.

The gene was expressed under the control of a triple promoter system (as described in WO 99/43835), consisting of the promoters from Bacillus licheniformis alpha-amylase gene (amyL), Bacillus amyloliquefaciens alpha-amylase gene (amyQ), and the Bacillus thuringiensis crylllA promoter including stabilizing sequence.

The gene was expressed with a Bacillus clausii secretion signal (encoding the following amino acid sequence: MKKPLGKIVASTALLISVAFSSSIASA (SEQ ID NO: 28) replacing the native se- cretion signal. Furthermore the expression construct results in the addition of an amino terminal poly histidine purification tag on the natural mature protein allowing for enzyme purification through immobilized metal ion affinity chromatography.

The SOE-PCR product was transformed into Bacillus subtilis and integrated in the chromosome by homologous recombination into the pectate lyase locus. Subsequently one recombinant Ba- cillus subtilis clone containing the respective galactanase expression construct was selected and was cultivated on a rotary shaking table in 500 ml baffled Erlenmeyer flasks each contain- ing 100 ml rich starch based media. After 3-5 days cultivation time at 30 °C to 37°C, enzyme containing supernatants were harvested by centrifugation and the enzymes were purified by immobilized metal affinity chromatography. Example 2: Purification of GH53 galactanases from Cohnella sp-60555 (SEQ ID NO: 4), Cohnella xylanilytica (SEQ ID NO: 8), Paenibacillus sp-18179 (SEQ ID NO: 12), Paenibacil- lus peoriae (SEQ ID NO: 16), Paenibacillus xylanexedens (SEQ ID NO: 20) and Cohnella laeviribosi (SEQ ID NO: 24)

The pH of the supernatant from example 1 was adjusted to pH 8, filtrated through a 0.2mM filter, and then applied to a 5 ml HisTrap™ excel column(GE Healthcare Life Sciences, Pittsburgh, USA). Prior to loading, the column had been equilibrated in 5 column volumes (CV) of 50 mM Tris/HCI pH 8. In order to remove unbound material, the column was washed with 8 CV of 50 mM Tris/HCI pH 8, and elution of the target was obtained with 50 mM HEPES pH 7 + 10mM im- idazole. The eluted protein was desalted on a HiPrep™ 26/10 desalting column (GE Healthcare Life Sciences, Pittsburgh, USA)., equilibrated using 3 CV of 50 mM HEPES pH 7 + 100 mM NaCI. This buffer was also used for elution of the target, and the flow rate was 10 ml/min. Rele- vant fractions were selected and pooled based on the chromatogram and SDS-PAGE analysis.

Example 3: Determination of Galactanase activity

Galactanase activity was determined by reducing ends using the colorimetric assay developed by Lever (Analytical Biochemistry 47, 273-279, 1972). Galactanase solubilizes sugars with re- ducing ends which react with the PAH BAH (Sigma H-9882) generating an increase of color which is proportional to the enzyme activity under the conditions used in the assay.

Soybean meal was filled with a solid dispenser into 96 well format plates. The weight was measured before and after addition of soybean meal and the substrate amount per well was es- timated assuming equal distribution along the plate.

50 pi enzyme sample and 50 mI activity buffer were transferred to the PCR-plate and mixed. The plate was incubated in a PCR machine at 40°C for 1 hour. Immediately after this the tempera- ture was lowered to 10°C.

75 mI of the freshly prepared PAHBAH solution was added to each well, mixed, and 75 mI of the mix was discarded. The mix was then incubated for 10 min at 95°C, and then for 1 min at 10°C.

150 mI were transferred manually to a new 96 MTP, and absorbance was measured at 405 nm.

Activity buffer: 100 mM acetate, 100 mM MES, (100 mM Glycine) in 0.01 % Triton X100, 1 mM CaCh, pH 5.

PAHBAH solution: 15 mg/ml PAHBAH, potassium sodium tartrate tetrahydrate (50g/L), NaOH (20g/L)

The enzyme samples were diluted to a concentration of 0.6 ppm (final enzyme concentration).

Conclusion: Galactanase from Cohnella sp-60555 (SEQ ID NO: 4), Galactanase from Cohnella xylanilytica (SEQ ID NO: 8), Galactanase from Paenibacillus sp-18179 (SEQ ID NO: 12), Galac- tanase from Paenibacillus peoriae (SEQ ID NO: 16), Galactanase from Paenibacillus xylanexe- dens (SEQ ID NO: 20) and Galactanase from Cohnella laeviribosi (SEQ ID NO: 24) all show higher activity on soybean meal at pH 5 compared to the benchmark Galactanases from H. in- solens and B. licheniformis.

Example 4: Determination of Galactanase activity

50 pi enzyme sample and 50 mI activity buffer were transferred to the PCR-plate and mixed. The plate was incubated in PCR machine at 40°C for 2 hours. Immediately thereafter the tempera- ture was lowered to 10°C. 75 pi of the freshly prepared PAHBAH solution was added to each well, mixed, and 75 mI of the mix was discarded. The mix was then incubated for 10 min at 95°C, then 1 min at 10°C.

150mI were transferred manually to a new 96 MTP, and absorbance was measured at 405nm.

Activity buffer: 100mM acetate, 100mM MES, (100mM Glycine) in 0.01% Triton X100, 1 mM CaCI₂, pH 6.5

Conclusion: Galactanase from Cohnella sp-60555 shows higher activity on soybean meal at pH 6.5 at all tested enzyme dosages compared to the benchmark Galactanases from H. insolens and B. licheniformis.

Example 5: Cloning of GH53 galactanase from Xanthomonas cynarae (SEQ ID NO: 32),

Galactanase from Xanthomonas cynarae (Seq ID No: 32) was cloned according to the proce- dure described in example 1.

Example 6: Purification of GH53 galactanase from Xanthomonas cynarae (SEQ ID NO: 32) Galactanase from Xanthomonas cynarae (Seq ID No: 32) was purified according to the proce- dure described in example 2.

Example 7: Determination of Galactanase activity

Galactanase activity was determined by reducing ends using the colorimetric assay adapted from Fridemann, Weber and Witt (Analytical Biochemistry 4, 358-377, 1962). Galactanase solu- bilizes sugars with reducing ends which react with the Potassium Ferricyanide (K₃Fe(CN)₆) (Sigma 244023) generating a decrease of color which is proportional to the enzyme activity un- der the conditions used in the assay.

Soybean meal dispersion diluted in Acetate buffer (pH 6.0) was filled with a liquid dispenser into 2ml_ microtubes. The dispersion was kept under agitation and it was assumed equal distribution along the tubes.

100 pi or 200 mI enzyme sample were transferred to the microtubes. The tubes were incubated in Thermomixer machine at 37°C for 2 hours. Immediately thereafter the temperature was low- ered to 10°C.

750 mI of the prepared Ferricyanide (KsFe(CN)₆) solution was added to new empty 2 ml. tubes and 95 mI of the hydrolyzed soybean meal dispersion added to each tube and mixed. The mix was then incubated for 12 min at 96°C, then cooled to room temperature.

250mI were transferred manually to a new 96 MTP, and absorbance was measured at 420nm.

Acetate buffer: 100mM acetate, pH 6.0

Ferricyanide solution: 1.2 g/l Potassium Ferricyanide, Sodium Carbonate (20 g/L)

Conclusion: Galactanase from Cohnella sp-60555, Cohnella laeviribosi, Paenibacillus xylanex- edens, Cohnella xylanilytica, Xanthomonas cynarae and Paenibacillus sp-18179 show higher activity on soybean meal at pH 6.0 at all tested enzyme dosages compared to the benchmark Galactanases from H. insolens.

Example 8: Determination of Galactanase activity

Soybean meal dispersion diluted in Acetate buffer (pH 4.5) was filled with a liquid dispenser into 2ml_ microtubes. The dispersion was kept under agitation and it was assumed equal distribution along the tubes.

100 pi or 200 mI enzyme sample were transferred to the microtubes. The tubes were incubated in Thermomixer machine at 50°C for 1 hour. Immediately thereafter the temperature was low- ered to 10°C.

Acetate buffer: 100mM acetate, pH 4.5

Conclusion: Galactanase from Cohnella sp-60555, Cohnella xylanilytica and Paenibacillus sp- 18179 show higher activity on soybean meal at pH 4.5 at tested enzyme dosage compared to the benchmark Galactanases from H. insolens.

Example 9: Determination of Galactanase activity

Galactanase activity was determined by nitrogen/protein quantification of hydrolyzed material using Dumas method (Leco® Instrument). Galactanase solubilizes sugars which are removed by alcoholic extraction, therefore an increase in the final nitrogen content is expected compared to the control treatment without enzyme.

Soybean meal slurry with adjusted pH 4.5 was filled manually into 50 ml. tubes.

Desired amounts of enzyme samples were transferred to the tubes. The tubes were incubated in rotisserie oven machine at 50°C for 4 hours. Immediately thereafter the desired amount of absolute ethanol was added.

The tubes were incubated in rotisserie oven machine at 60°C for 15 minutes and then centri- fuged at 15°C for 10 minutes. The liquid fraction was removed and the procedure was repeated to the complete removal of the solubilized sugars.

The material was dried at 60°C overnight and then milled into fine powder. It was then analyzed for nitrogen content using Leco® Instrument (standard combustion conditions: 900°C furnace temperature). By using the protein factor of 6.25, the results were expressed in Protein (%).

Conclusion: Galactanase from Cohnella sp-60555 shows higher activity on soybean meal at pH 4.5 at tested enzyme dosage compared to the benchmark Galactanases from H. insolens, which is observed after a higher solubilization of sugars after hydrolysis of SBM.

Claims

i) providing a soy-based material and

87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 23; (g) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 31 ;

(k) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 9; (l) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 13;

(p) a variant of SEQ ID NO: 7 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(t) a variant of SEQ ID NO: 23 wherein the variant has galactanase activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;

(v) a polypeptide comprising the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (I), (m), (n), (o), (p), (q), (r), (s), (t), or (u) and a N-terminal and/or C- terminal His-tag and/or HQ-tag; and

(w) a fragment of the polypeptide of (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (I), (m), (n), (o), (p), (q), (r), (s), (t), (u) or (v) having galactanase activity and having at least 90% of the length of the mature polypeptide, wherein the polypeptide having galactanase activity has at least 60%, such as at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95% or at least 100% of the galactanase activity of the polypeptide of SEQ ID NO: 3.

2. The method according to claim 1 , wherein the soy-based material is selected from the group consisting of dehulled full-fat flakes, full-fat soybean meal, defatted flakes, white flakes, full-fat flakes and soybean meal.

3. The method according to any of the previous claims, wherein the soy-based material is in an aqueous solution.

4. The method according to any of the previous claims, wherein the soy-based material has a dry-matter content of from 20-40% (w/w).

5. The method according to any of claims 1 to 3, wherein the soy-based material has a dry- matter content of from 40-60% (w/w).

6. The method according to any of the previous claims, wherein galactanase treated soy-based material has an increased protein content (compared to soy-based material that has not been treated with galactanase).

7. The method according to any of the previous claims, wherein galactanase treated soy-based material has an increased NDF digestibility (compared to soy-based material that has not been treated with galactanase).

8. The method according to any of the previous claims, wherein step ii) comprises galactanase treatment of the soy-based material at pH between pH 3 and 6, such as between pH 3 and 4, for example between pH 4 and 5, such as pH 5 and 6, or any combination of these intervals.

9. The method according to any of the previous claims, wherein step ii) comprises galactanase treatment of the soy-based material at a temperature between 40°C and 80°C such as between 50°C and 70°C.

10. The method according to any of the previous claims, wherein step ii) comprises galactanase treatment of the soy-based material at a temperature between 15°C and 40°C such as between 20°C and 30°C.

1 1. The method according to any of the previous claims, wherein the method results in production of soybean meal, fermented soybean meal or soy protein concentrate.

12. The method according to any of the previous claims, wherein the polypeptide comprises or consists of amino acids 1 to 316 of SEQ ID NO: 2, amino acids 1 to 316 of SEQ ID NO: 3, ami- no acids 1 to 324 of SEQ ID NO: 4, amino acids 1 to 318 of SEQ ID NO: 6, amino acids 1 to 318 of SEQ ID NO: 7, amino acids 1 to 326 of SEQ ID NO: 8, amino acids 1 to 516 of SEQ ID NO: 10, amino acids 1 to 516 of SEQ ID NO: 1 1 , amino acids 1 to 524 of SEQ ID NO: 12, amino ac- ids 1 to 317 of SEQ ID NO: 14, amino acids 1 to 317 of SEQ ID NO: 15, amino acids 1 to 325 of SEQ ID NO: 16, amino acids 1 to 316 of SEQ ID NO: 18, amino acids 1 to 316 of SEQ ID NO: 19, amino acids 1 to 324 of SEQ ID NO: 20, amino acids 1 to 316 of SEQ ID NO: 22, amino ac- ids 1 to 316 of SEQ ID NO: 23, amino acids 1 to 324 of SEQ ID NO: 24, amino acids 1 to 307 of SEQ ID NO: 30, amino acids 1 to 307 of SEQ ID NO: 31 or amino acids 1 to 315 of SEQ ID NO: 32.

13. An animal feed or animal feed additive comprising the galactanase treated soy-based material obtained in the method according to any of claims 1 to 12.

14. Use of an isolated polypeptide having galactanase activity for improving the nutritional value of soy-based material, wherein the polypeptide is selected from the group consisting of:

(d) a polypeptide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the polypeptide of SEQ ID NO: 15;

(i) the mature polypeptide coding sequence of SEQ ID NO: 1 or the cDNA sequence thereof, (ii) the mature polypeptide coding sequence of SEQ ID NO: 5 or the cDNA sequence thereof,

(m) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 17; (n) a polypeptide encoded by a polynucleotide having at least 80%, e.g., at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 21 ;

wherein the polypeptide having galactanase activity has at least 60%, such as at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95% or at least 100% of the galactanase activity of the polypeptide of SEQ ID NO: 3.

15. The use according to claim 14, wherein the polypeptide comprises or consists of amino ac- ids 1 to 316 of SEQ ID NO: 2, amino acids 1 to 316 of SEQ ID NO: 3, amino acids 1 to 324 of SEQ ID NO: 4, amino acids 1 to 318 of SEQ ID NO: 6, amino acids 1 to 318 of SEQ ID NO: 7, amino acids 1 to 326 of SEQ ID NO: 8, amino acids 1 to 516 of SEQ ID NO: 10, amino acids 1 to 516 of SEQ ID NO: 11 , amino acids 1 to 524 of SEQ ID NO: 12, amino acids 1 to 317 of SEQ ID NO: 14, amino acids 1 to 317 of SEQ ID NO: 15, amino acids 1 to 325 of SEQ ID NO: 16, amino acids 1 to 316 of SEQ ID NO: 18, amino acids 1 to 316 of SEQ ID NO: 19, amino acids 1 to 324 of SEQ ID NO: 20, amino acids 1 to 316 of SEQ ID NO: 22, amino acids 1 to 316 of SEQ ID NO: 23, amino acids 1 to 324 of SEQ ID NO: 24, amino acids 1 to 307 of SEQ ID NO: 30, amino acids 1 to 307 of SEQ ID NO: 31 or amino acids 1 to 315 of SEQ ID NO: 32.