JP2001061473A - New polygalacturonase - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain the subject new enzyme derived from Acremonium cellulolyticus as a kind of filamentous fungus and capable of hydrolyzing polygalacturonic acid through endo mechanism, and useful for food quality improvement in food processing, feed efficiency improvement, scouring and feel improvement of cotton, etc. SOLUTION: This enzyme is a new polygalacturonase 1 having an N- terminated amino acid sequence of the formula which is derived from Acremonium cellulolyticus as a kind of filamentous fungus and capable of hydrolyzing polygalacturonic acid through endo mechanism. Furthermore, the new enzyme has the following characteristics: for reducing sugar formation activity with polygalacturonic aid as substrate, optimum pH is 4.5, being stable at pH 3.5-8.5 (4 deg.C, 24 h); for reducing sugar formation activity with polygalacturonic aid as substrate, action optimum temperature is 50 deg.C, being stable at <=40 deg.C (pH 4.5, 10 min); isoelectric point is pI 6.9 (polyacrylamide gel isoelectric point electrophoresis); and molecular weight is 42,000 (SDS- polyacrylamide gel electrophoresis). The enzyme is useful for improving both the quality and yield of food in its processing, improving feed efficiency and silage quality, and for scouring cotton, etc.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to pectinases, and more particularly to a novel polygalacturonase derived from the filamentous fungus Acremonium cellulolyticus.

[0002]

BACKGROUND OF THE INVENTION Pectin is a major constituent of higher plant intercellular material and is a naturally occurring substance in a wide range and in large quantities. Pectin is expressed in units of galacturonic acid.
The basic skeleton is a high-molecular polysaccharide, polygalacturonic acid, which is linearly polymerized by -1,4-galacturonoside bonds, and the carboxylic acid at the 6-position is partially methylated in nature. Further, L-rhamnose is found in the main chain, and L-arabinose, D-galactose, D-xylose, and the like are found in the side chains. In addition, other components, especially hemicelluloses, furthermore, cellulose, lignin and the like are complexly combined or interact with each other to form a plant tissue.

[0003] Pectinase is a general term for a group of enzymes that catalyze the enzyme reaction system that degrades pectin. Polygalacturonase, which hydrolyzes the α-1,4 bond of pectin or polygalacturonic acid, is mainly called β-elimination reaction. Pectin lyase and polygalacturonic acid lyase, which degrade chains, and pectin methylesterase, which hydrolyzes the methyl ester of pectin, have been reported to date on an extremely large number of pectinases. Pectinase is an enzyme positioned at the center of a group of enzymes that disrupt plant tissues, and is particularly important in maceration and plant cell engineering to release plant cells without destroying them. Furthermore, plant cells can be completely destroyed in cooperation with hemicellulases such as xylanase, galactanase, arabinase, and mannanase, cellulases, and lignin-degrading enzymes. In application fields, clarification of fruit juice,
It is expected to improve filterability, yield, food processing such as pureing vegetable fruits, improve feed efficiency by adding to feed, and promote fermentation and quality by adding to silage.

[0004] Speaking of enzymes produced by the filamentous fungus Acremonium cellulolyticus , cellulase is characterized by a strong saccharification activity, and its usefulness in feed and silage applications has been reported ( For example, JP-A-4-117244, JP-A-7-236431
No.). In addition, cellulase components contained therein have been reported (for example, see Agric. Biol. Chem., 5).
2, 2493〜2501 (1988), ibid.53, 3359〜3360 (1989),
54, 309-317 (1990)). In addition to cellulase, xylanase and pectinase enzyme activities are present, and it has been reported that pectin of legume alfalfa is degraded (for example, Agric. Biol. Chem.,
51, 65-74 (1987), Journal 51, 3207-3213 (1987), Grass
land Science, 42, 159-162 (1996)). Furthermore, partial purification of some pectinase was also attempted (Grass
land Science, 42, 343-347 (1997)). However, the purification of pectinase is insufficient, and thus the amino acid sequence and properties of the enzyme have not been studied, and it has not been possible to use it industrially effectively.

[0005]

SUMMARY OF THE INVENTION Accordingly, the present inventors
Various enzymes constituting the pectinase system derived from Acremonium cellulolyticus were fractionated, purified, and studied intensively. As a result, an active ingredient in the enzyme group was found, and the present invention was completed based on this finding. An object of the present invention is to provide a novel polygalacturonase that is effective for applications such as food processing, feed, and silage.

[0006]

Means for Solving the Problems The present invention according to claim 1 is polygalacturonase 1 derived from a filamentous fungus Acremonium cellulolyticus and having the following properties. (a) Action: This enzyme hydrolyzes polygalacturonic acid by endo mechanism. (b) Substrate specificity: This enzyme acts on polygalacturonic acid. (c) Optimum pH and stable pH range: In the reducing sugar-forming activity using polygalacturonic acid as a substrate, the optimum pH is 4.5, and the pH is stable in the range of 3.5 to 8.5 (4 ° C., 24 hours). . (d) Optimum action temperature: 50 ° C. for reducing sugar-forming activity using polygalacturonic acid as a substrate. (e) Temperature stability: stable at 45 ° C or less (pH 4.5,
10 minutes). (f) Isoelectric point: pI 6.9 (by polyacrylamide gel isoelectric focusing) (g) Molecular weight: 42,000 (by SDS-polyacrylamide gel electrophoresis) (h) Specific activity: 208.3 units / mg protein (polygalacturonic acid-reducing sugar-forming activity) (i) N-terminal amino acid sequence: has the sequence shown in SEQ ID NO: 1 in the sequence listing.

The present invention according to claim 2 is polygalacturonase 2 derived from the filamentous fungus Acremonium cellulolyticus and having the following properties. (a) Action: This enzyme hydrolyzes polygalacturonic acid by endo mechanism. (b) Substrate specificity: This enzyme acts on polygalacturonic acid. (c) Optimum pH and stable pH range: In the reducing sugar-forming activity using polygalacturonic acid as a substrate, the optimum pH is 4.5, and the pH is stable in the range of 2.5 to 12.0 (4 ° C., 24 hours). . (d) Optimum action temperature: 50 ° C. for reducing sugar-forming activity using polygalacturonic acid as a substrate. (e) Temperature stability: stable at 50 ° C or less (pH 4.5,
10 minutes). (f) Isoelectric point: pI4.1 (by polyacrylamide gel isoelectric focusing) (g) Molecular weight: 41,000 (by SDS-polyacrylamide gel electrophoresis) (h) Specific activity: 56.1 units / mg protein (polygalacturonic acid-reducing sugar-forming activity) (i) N-terminal amino acid sequence: has the sequence shown in SEQ ID NO: 2 in the sequence listing.

[0008] The present invention according to claim 3 is based on claim 1.
An enzyme for enhancing and utilizing polygalacturonase 1 according to claim 2 and / or polygalacturonase 2 according to claim 2 for hydrolyzing pectin to utilize plant tissues or plant tissue-derived products. It is a preparation.

Further, the present invention according to claim 4 is characterized in that the enzyme preparation according to claim 3 further comprises cellulase, xylanase, protease, galactanase, arabinase, mannanase, rhamnogalacturonase, pectin methylesterase, pectin. An enzyme preparation comprising one or more components of lyase and polygalacturonic acid lyase.

[0010]

BEST MODE FOR CARRYING OUT THE INVENTION The pectinase of the present invention is an active ingredient contained in a pectinase system produced by Acremonium cellulolyticus. Hereinafter, the present invention will be described in detail. Microorganisms producing the pectinase system containing the polygalacturonase of the present invention include Acremonium cellulolyticus strain Y-94 (FERM BP-5826), Acremonium cellulolyticus TN strain (FERM BP-685), and the like. Production of pectinase by the present bacterium may be carried out, for example, according to the methods described in JP-B-60-43954 and JP-B-63-63197. After the cultivation of the microorganism, the supernatant obtained by removing the culture by centrifugation or the like can be used as a crude enzyme.However, usually, the supernatant is concentrated by ultrafiltration, etc. A concentrated enzyme is obtained by adding an agent or the like, or a powdered enzyme is obtained by a spray drying method after concentration.

The polygalacturonase of the present invention can be obtained by partially or highly purifying these concentrated or powdered enzymes as required. Examples of the purification method include a conventional method, ie, a salting-out method using ammonium sulfate or the like; an organic solvent precipitation method using an alcohol or the like; a membrane separation method; , Alone or in combination as appropriate.

The properties of the highly purified polygalacturonase (polygalacturonase 1, polygalacturonase 2) obtained by the present invention are as follows.
This enzyme is a novel enzyme not known in the literature.

[0013] Action of polygalacturonase 1 (a): This enzyme hydrolyzes polygalacturonic acid by endo mechanism. (b) Substrate specificity: This enzyme acts on polygalacturonic acid. (c) Optimum pH and stable pH range: In the reducing sugar-forming activity using polygalacturonic acid as a substrate, the optimum pH is 4.5, and the pH is stable in the range of 3.5 to 8.5 (4 ° C., 24 hours). . (d) Optimum action temperature: 50 ° C. for reducing sugar-forming activity using polygalacturonic acid as a substrate. (e) Temperature stability: stable at 45 ° C or less (pH 4.5,
10 minutes). (f) Isoelectric point: pI 6.9 (by polyacrylamide gel isoelectric focusing) (g) Molecular weight: 42,000 (by SDS-polyacrylamide gel electrophoresis) (h) Specific activity: 208.3 units / mg protein (polygalacturonic acid-reducing sugar-forming activity) (i) N-terminal amino acid sequence: has the sequence shown in SEQ ID NO: 1 in the sequence listing.

Polygalacturonase 2 (a) action: This enzyme hydrolyzes polygalacturonic acid by endo-mechanism. (b) Substrate specificity: This enzyme acts on polygalacturonic acid. (c) Optimum pH and stable pH range: In the reducing sugar-forming activity using polygalacturonic acid as a substrate, the optimum pH is 4.5, and the pH is stable in the range of 2.5 to 12.0 (4 ° C., 24 hours). . (d) Optimum action temperature: 50 ° C. for reducing sugar-forming activity using polygalacturonic acid as a substrate. (e) Temperature stability: stable at 50 ° C or less (pH 4.5,
10 minutes). (f) Isoelectric point: pI4.1 (by polyacrylamide gel isoelectric focusing) (g) Molecular weight: 41,000 (by SDS-polyacrylamide gel electrophoresis) (h) Specific activity: 56.1 units / mg protein (polygalacturonic acid-reducing sugar-forming activity) (i) N-terminal amino acid sequence: has the sequence shown in SEQ ID NO: 2 in the sequence listing.

Here, the method for measuring the activity of the enzyme and the definition of one unit are as follows.・ Polygalacturonase reducing sugar-forming activity At pH 4.5, 30 ° C, an enzyme is allowed to act on a 0.025% polygalacturonic acid (orange origin) suspension, and 1 μmo per minute
The amount of the enzyme that produces a reducing sugar corresponding to 1 galacturonic acid was defined as 1 unit.

The polygalacturonase obtained by the present invention has excellent pectin-degrading performance, and is suitable for processing and improving the utilization of pectin-containing plant tissues or plant tissue-derived products. It is the invention according to claim 3 that provides such an enzyme preparation. That is, claim 3
The present invention according to the present invention relates to a plant tissue or a plant tissue by hydrolyzing pectin, which contains polygalacturonase 1 according to claim 1 and / or polygalacturonase 2 according to claim 2 in an enhanced manner. It is an enzyme preparation for processing using a derived material.

The present enzymes (polygalacturonase 1 and polygalacturonase 2) can be used alone or in combination as described in claim 3, respectively. As described in item 4,
In addition to these, cellulase, xylanase, protease, galactanase, arabinase, mannanase,
It can also be used as an enzyme preparation in which any one or more components of rhamnogalacturonase, pectin methylesterase, pectin lyase and polygalacturonic acid lyase are combined. In the case of the present invention according to claim 4, claim 3 , The efficiency can be further enhanced as compared with the present invention.

The polygalacturonase or enzyme preparation described in any one of claims 1 to 4 can be used for processing and processing of pectin-containing plant tissues or plant tissue-derived products. Here, the plant tissue or the plant tissue-derived product can be applied to all plant tissues. In particular, vegetables and fruits are targeted in the food field, and can be applied to processed fruit juice, puree, paste, pomace, and extracted pomace. In the field of feed, alfalfa containing relatively pectin is targeted for silage use,
Grains such as defatted soybeans and cottonseed oil lees are also targeted for feed addition for the same reasons. The polygalacturonase or the enzyme preparation according to any one of claims 1 to 4 can be added to livestock feed to improve feed efficiency. In the field of fibers, cotton and hemp are targeted, and can be used for scouring or improving texture.

The enzyme and the enzyme preparation can be used under the conditions such as pH and temperature generally used in the above-mentioned application fields.
A temperature range of 10-65 ° C is suitable. The amount of enzyme used is
It is possible to obtain a sufficient effect if adjustment is made so that each purpose is achieved with time.

[0020]

Next, the present invention will be described in detail with reference to examples, but the scope of the present invention is not limited to these examples.

Example 1 (Preparation of bulk enzyme powder) In order to obtain a bulk pectinase derived from a microorganism of the genus Acremonium, the microorganism was cultured by the following method. The medium used was a medium having the following composition and heat-sterilized by a conventional method.

[Medium composition] Cotton seed oil lees 2%, cellulose 2
%, Dipotassium hydrogen phosphate 1.2%, bactopeptone 1
%, 0.6% potassium nitrate, 0.2% urea, 0.4% potassium chloride.
16%, magnesium sulfate / heptahydrate 0.12%, zinc sulfate / heptahydrate 0.001%, manganese sulfate / heptahydrate 0.001%
%, Copper sulfate pentahydrate 0.001% (pH 4.0).

In 500 ml of the above medium, add acremonium
After inoculating Cellulolyticus TN strain (FERM BP-685),
The cells were cultured with stirring at 0 ° C. for 48 hours. Next, this culture solution was scaled up to 15 L as a seed, and further scale-up was continued to finally adjust the volume of the culture solution in the 600 L tank to 300 L, followed by aeration and stirring culture for 7 days. The resulting culture was filtered with a filter press, concentrated to 15 L by ultrafiltration, lactose (2 kg) was added, and powdered by spray drying. The bulk pectinase obtained by this method was 5.0 kg.

Example 2 (Purification of polygalacturonase) The bulk powder obtained in Example 1 was dissolved in an acetate buffer (pH 5.5), and impurities were removed by high-speed cooling centrifugation. The obtained supernatant was purified by the following method as a starting material for enzyme purification.

Strongly basic anion exchange chromatography: QAE-Toyopearl 550C (Tosoh Corporation) was used to adsorb the supernatant, and NaCl was added to acetate buffer (0.02M, pH 5.5).
Was contained in each of 0 M, 0.15 M and 0.5 M, and stepwise elution was carried out, and polygalacturonase active fractions (I and IV fractions) eluted with 0 M and 0.5 M NaCl were collected.

Strongly acidic ion exchange chromatography:
The above fraction I was adsorbed on Mono S (Pharmacia) with acetate buffer (0.02 M, pH 4.5), and NaCl was contained in acetate buffer (0.02 M, pH 4.5) from 0 M to 0.2 M. A linear gradient elution was performed, and a fraction I-4 (polygalacturonase 1) showing polygalacturonase activity was collected.

Strongly basic ion exchange chromatography: The above fractionated IV fraction was adsorbed to Mono Q (Pharmacia) with an acetate buffer (0.02 M, pH 5.5), and the acetate buffer (0.02 M, pH 5. A linear gradient elution containing 0 M to 0.5 M of NaCl in 5) was carried out, and fraction IV-2 (polygalacturonase 2) showing polygalacturonase activity was obtained.
Was collected.

Example 3 (Substrate Specificity and Action Characteristics) The activity of the purified polygalacturonase (polygalacturonase 1, 2) obtained in Example 2 on various substrates was examined.
Two enzymes were separately acted on pectin and polygalacturonic acid of various origins, and the saccharides produced over time were separated and detected by thin-layer chromatography. It was found that they mainly produce sugar, and that both enzymes act on the substrate in an endo mechanism.

Example 4 The optimal pH of polygalacturonase 1 and 2 obtained in Example 2 at 30 ° C. are shown in FIGS. 1 and 2, respectively. These enzymes showed maximum activity in McIlvaine buffer at pH 4.5. The pH stability of these enzymes at 4 ° C. is shown in FIGS. 3 and 4, respectively, and the pH stability at 45 ° C. is shown in FIGS. 5 and 6, respectively. 3 to 6 indicate McIlvaine buffer, and □ indicates Britton.
& Robinson buffer is shown.

Example 5 In order to examine the optimal temperature of action of polygalacturonase 1 and 2 obtained in Example 2, the reducing sugar-forming activity using polygalacturonic acid (origin orange) as a substrate was measured. As shown in FIG.
(PH 4.5). Next, the temperature stability of these enzymes was measured under conditions of pH 4.5 and 10 minutes, and as shown in FIGS.
It was stable below ℃.

Example 6 To determine the molecular weight of polygalacturonase 1 and 2 obtained in Example 2, SDS-polyacrylamide gel electrophoresis was performed. That is, a 12.5% gel was applied at a constant current of 20 mA for 80 minutes at room temperature. as a result,
The molecular weight of polygalacturonase 1 is about 42,000,
The molecular weight of polygalacturonase 2 was calculated to be about 41,000.

The molecular weight of the standard sample used in this test is as follows. Phosphorylase b: 97,000 Serum albumin: 66,000 Ovalbumin: 45,000 Carbonic anhydrase: 31,000 Trypsin inhibitor: 21, 500 Lysozyme: 14,400

Example 7 The N-terminal amino acid sequence of each of polygalacturonase 1 and 2 obtained in Example 2 was determined. First, 8% Gel
After electrophoretic separation of each sample using SDS-PAGE mini (manufactured by Tefco), the sample was transferred to a mini-problot membrane (manufactured by PerkinElmer) using Horizon Blot (manufactured by Atto). Coomassie Brilliant Blue R-25
0 (manufactured by Nacalai Tesque), washed with 50% methanol and air-dried. From this, a portion where a protein having a molecular weight of 42,000 or 41,000 was blotted was cut out, and a protein sequencer: Procise 491 was cut out.
(Perkin Elmer) to determine the N-terminal amino acid, and determined the N-terminal amino acid sequence of polygalacturonase 1 at 17 residues and the N-terminal amino acid sequence of polygalacturonase 2 at 15 residues. . The obtained sequence was as shown in SEQ ID NOs: 1 and 2 in the sequence listing.

[0034]

The enzyme polygalacturonase of the present invention comprises
It is the main component of pectinase from Acremonium cellulolyticus, and its properties have been elucidated for the first time. The enzyme (polygalacturonase 1, 2) is effective in various uses such as quality improvement in food processing, yield improvement, feed efficiency improvement, silage quality improvement, cotton and hemp scouring, and texture improvement.

[Sequence List] SEQUENCE LISTING <110> Meiji Seika Co., Ltd. MEIJI SEIKA KAISHA LTD. <110> Director of the Industrial Technology Director 210> 1 <211> 17 <212> PRT <213> Acremonium cellulolyticus <400> 1 Ala Ala Ser Tyr Thr Phe Ser Gly Ser Asn Gly Ala Ala Cys Ala Cys Lys 1 5 10 15 17 <210> 2 <211> 15 <212> PRT <213> Acremonium cellulolyticus <400> 2 Ala Thr Ser Asn Thr Phe Asp Gly Asp Asp Gly Ala Ala Cys Ala 1 5 10 15

[Brief description of the drawings]

FIG. 1 shows the polygalacturonase 1 obtained in Example 2
It is a figure which shows the optimal pH in 0 degreeC.

FIG. 2 shows a graph of polygalacturonase 2 obtained in Example 2.
It is a figure which shows the optimal pH in 0 degreeC.

FIG. 3 shows 4 of polygalacturonase 1 obtained in Example 2.
FIG. 4 is a diagram showing pH stability at ° C.

FIG. 4 shows a diagram of 4 of polygalacturonase 2 obtained in Example 2.
FIG. 4 is a diagram showing pH stability at ° C.

FIG. 5 shows 4 of polygalacturonase 1 obtained in Example 2.
It is a figure which shows pH stability in 5 degreeC.

6 shows 4 of polygalacturonase 2 obtained in Example 2. FIG.
It is a figure which shows pH stability in 5 degreeC.

FIG. 7 is a view showing an optimum temperature of polygalacturonase 1 obtained in Example 2.

FIG. 8 is a graph showing the optimum temperature of polygalacturonase 2 obtained in Example 2.

FIG. 9 is a graph showing the temperature stability of polygalacturonase 1 obtained in Example 2.

FIG. 10 is a graph showing the temperature stability of polygalacturonase 2 obtained in Example 2.

[Explanation of symbols]

3 to 6 indicate McIlvaine buffer, and □ indicates Britto
n & Robinson buffer.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) C12P 21/02 C12N 15/00 ZNAA // (C12N 1/14 C12R 1: 645) (C12N 15/09 ZNA (C12P 1: 645) (C12P 21/02 C12R 1: 645) (72) Takanori Nihei Inventor 739-1 Otani, Oya, Shizuoka City, Shizuoka Prefecture C102 (72) Inventor Sohei Nojiri 5 Chiyoda, Sakado City, Saitama Prefecture -3-1 Meiji Seika Co., Ltd. Biological Science Laboratory Co., Ltd. (72) Inventor Asayuki Fukasawa 5-3-1 Meiji Seika Co., Ltd. Biological Science Laboratories Co., Ltd. (72) Inventor Toshiaki Kono Chiyoda, Sakado City, Saitama Prefecture 5-3-1 Meiji Seika Co., Ltd. Biological Science Laboratory Co., Ltd. (72) Inventor Rin Yamabe 1-1-3 Higashi, Tsukuba, Ibaraki Pref. Work Institute of Technology in the F-term (reference) 2B150 AB01 DD11 DF11 4B024 AA05 AA20 BA12 4B050 CC01 CC08 DD03 FF11E LL02 4B064 AG01 BA14 CA05 CE11 DA10 4B065 AA58X AC14 CA31 CA41

Claims (4)

[Claims] 1. A polygalacturonase 1 derived from a filamentous fungus Acremonium cellulolyticus and having the following properties: (a) Action: This enzyme hydrolyzes polygalacturonic acid by endo mechanism. (b) Substrate specificity: This enzyme acts on polygalacturonic acid. (c) Optimum pH and stable pH range: In the reducing sugar-forming activity using polygalacturonic acid as a substrate, the optimum pH is 4.5, and the pH is stable in the range of 3.5 to 8.5 (4 ° C., 24 hours). . (d) Optimum action temperature: 50 ° C. for reducing sugar-forming activity using polygalacturonic acid as a substrate. (e) Temperature stability: stable at 45 ° C or less (pH 4.5,
10 minutes). (f) Isoelectric point: pI 6.9 (by polyacrylamide gel isoelectric focusing) (g) Molecular weight: 42,000 (by SDS-polyacrylamide gel electrophoresis) (h) Specific activity: 208.3 units / mg protein (polygalacturonic acid-reducing sugar-forming activity) (i) N-terminal amino acid sequence: has the sequence shown in SEQ ID NO: 1 in the sequence listing. 2. A polygalacturonase 2 derived from the filamentous fungus Acremonium cellulolyticus and having the following properties: (a) Action: This enzyme hydrolyzes polygalacturonic acid by endo mechanism. (b) Substrate specificity: This enzyme acts on polygalacturonic acid. (c) Optimum pH and stable pH range: In the reducing sugar-forming activity using polygalacturonic acid as a substrate, the optimum pH is 4.5, and the pH is stable in the range of 2.5 to 12.0 (4 ° C., 24 hours). . (d) Optimum action temperature: 50 ° C. for reducing sugar-forming activity using polygalacturonic acid as a substrate. (e) Temperature stability: stable at 50 ° C or less (pH 4.5,
10 minutes). (f) Isoelectric point: pI4.1 (by polyacrylamide gel isoelectric focusing) (g) Molecular weight: 41,000 (by SDS-polyacrylamide gel electrophoresis) (h) Specific activity: 56.1 units / mg protein (polygalacturonic acid-reducing sugar-forming activity) (i) N-terminal amino acid sequence: has the sequence shown in SEQ ID NO: 2 in the sequence listing. 3. The polygalacturonase 1 according to claim 1, and / or the polygalacturonase 2 according to claim 2.
An enzyme preparation for processing a plant tissue or a plant tissue-derived product by hydrolyzing pectin, the enzyme preparation containing the enzyme enhanced. 4. The enzyme preparation according to claim 3, further comprising any of cellulase, xylanase, protease, galactanase, arabinanase, mannanase, rhamnogalacturonase, pectin methylesterase, pectin lyase, and polygalacturonic acid lyase. An enzyme preparation comprising one or more components.