KR102612093B1 - Novel strain Aureobasidium pullulans RG4 having xylan degrading activity - Google Patents

Abstract

The present invention relates to a new strain of Aureobasidium pullulans RG4 having xylan decomposition activity, a method for decomposing polysaccharides using the same, a composition for decomposing polysaccharides, a method for producing disaccharides, and a composition for producing disaccharides.
According to the present invention, various polysaccharides, including xylan, can be decomposed very effectively through biological methods, and in particular, disaccharides can be effectively produced from xylan.

Description

Novel strain Aureobasidium pullulans RG4 having xylan degrading activity}

The present invention relates to a new strain of Aureobasidium pullulans RG4 having xylan decomposition activity, a method for decomposing polysaccharides using the same, a composition for decomposing polysaccharides, a method for producing disaccharides, and a composition for producing disaccharides.

Xylan is hemicellulose, one of the main components that accounts for about 20 to 40% of the cell walls of land plants, and is composed of arabinosyl, acetyl, and glucuronosyl. It has a complex structure composed of substituted β-1,4-linked xylose units, and can be hydrolyzed by various xylanases.

Xylanase is required in various fields such as the pulp-pre-bleaching process to remove hemicellulose from the pulp, promoting digestion of livestock feed, food and baking additives, fruit and vegetable processing, ethanol and xylitol production, and papermaking. Research on the characteristics of xylanase suitable for the field is required. In addition, xylooligosaccharide (XOS), a xylan hydrolysis product produced by xylanase, is known to have various physiological activities such as antibacterial, antioxidant, and anti-inflammatory, and is used in various fields such as food-pharmaceutical industry, agricultural-livestock industry, and cosmetics industry. It is expected that it can be used.

The decomposition method using a hydrolytic enzyme such as xylanase mentioned above has the advantage of being able to selectively and specifically hydrolyze polysaccharides to produce the target oligosaccharide. In particular, hydrolysis of plant biomass using microbial enzymes is accepted as an environmentally friendly alternative to chemical decomposition methods that produce harmful by-products. For this reason, many researchers are attempting to isolate new microorganisms that produce more suitable enzymes.

Accordingly, the present inventors sought to discover new microorganisms that could be used to biologically decompose (enzymatically) decompose xylan and produce specific decomposition products from xylan.

Christakopoulos P, Katapodis P, Kalogeris E, Kekos D, Macris BJ, Stamatis H, Skaltsa H. Antimicrobial activity of acidic xylo-oligosaccharides produced by families 10 and 11 endoxylanases. Int J Biol Macromol. 2003 Jan 15;31(4-5):171-5.

Therefore, the main purpose of the present invention is to provide a new microorganism that can be used to biologically decompose xylan and produce specific decomposition products from xylan.

Another object of the present invention is to provide a method of utilizing the microorganism.

Another object of the present invention is to provide a product utilizing the above microorganisms.

According to one aspect of the present invention, the present invention provides Aureobasidium pullulans RG4 strain deposited under accession number KACC 93361P.

According to another aspect of the present invention, the present invention provides a method for decomposing polysaccharides comprising contacting a culture of the strain or a purified product of the culture with a polysaccharide.

In the polysaccharide decomposition method of the present invention, the polysaccharide is preferably xylan, arabinoxylan, amylose, or xyloglucan.

According to another aspect of the present invention, the present invention provides a composition for decomposing polysaccharides comprising a culture of the above strain or a purified product of the above culture.

In the composition for decomposing polysaccharides of the present invention, the polysaccharide is preferably xylan, arabinoxylan, amylose, or xyloglucan.

According to another aspect of the present invention, the present invention provides a method for producing a disaccharide comprising contacting a culture of the strain or a purified product of the culture with xylan.

According to another aspect of the present invention, the present invention provides a composition for producing a disaccharide comprising a culture of the above strain or a purified product of the above culture.

According to the present invention, various polysaccharides, including xylan, can be decomposed very effectively through biological methods, and in particular, disaccharides can be effectively produced from xylan.

Figure 1 shows the NJ (Neighbor-Joining) phylogenetic tree analyzed based on the ITS-D1D2 sequence of the strain of the present invention.
Figure 2 shows a photograph taken through a microscope of the bacterial cells of the strain of the present invention grown on a solid medium (left) and a photograph taken of colonies of the strain of the present invention formed on a solid medium (right). In the colony photo on the right, the left side (marked '(top)') is a shot of the top of the medium, and the right side (marked '(bottom)') is a shot of the bottom of the medium. .
Figure 3 shows the results of a polysaccharide decomposition activity test of the strain of the present invention. (A), amylose decomposition activity test results; (B), xylan decomposition activity test results; (C), Arabinoxylan decomposition activity test results; (D), Xyloglucan decomposition activity test results.
Figure 4 shows the results of xylanase activity experiments according to culture time of the strain of the present invention.
Figure 5 shows the results of TLC analysis of the reaction product of the crude enzyme solution and xylan of the strain of the present invention. G, glucose; X2, xylobiose; X4, xylotetraose; X6, xylohexaose; R, reactant.
Figure 6 shows the microbial deposit of the strain of the present invention.

The new strain of the present invention, Aureobasidium pullulans ( Aureobasidium pullulans ) RG4, was isolated from rose flowers and deposited in the Korean Agricultural Culture Collection (KACC) of the National Institute of Agricultural Sciences under the accession number KACC 93361P.

The strain of the present invention has the ITS sequence of SEQ ID NO: 1 and the D1D2 sequence of SEQ ID NO: 2. The ITS sequence and D1D2 sequence are representative sequences used to identify fungi, and the ITS sequence of the strain of the present invention is different from the previously known ITS sequence of Aureobasidium pullulans.

The strain of the present invention is significantly different from Aureobasidium pullulans CBS 584.75, a representative existing Aureobasidium pullulans strain, in that it forms dark black colonies. In addition, the strain of the present invention can decompose polysaccharides such as xylan, arabinoxylan, amylose and xyloglucan, and in particular decomposes xylan to produce disaccharide, that is, a dimer (DP2) consisting of two monosaccharide molecules as the main product. can be produced

Aureobasidium pullulans is a yeast-like fungus that was first introduced in 1981 (Cooke, WB An Ecological Life History of Aureobasidium pullulans (De Bary) Arnaud. Mycopathol. Mycol. Appl. 1959, 12, 1??45.) , it is known to produce polysaccharides such as pullulan, anticancer substances such as liamocin, and antibacterial substances such as siderophores, and also produces useful enzymes. However, there has been no report on producing disaccharides by decomposing xylan like the strain of the present invention.

Therefore, the strain of the present invention differs from the existing Aureobasidium pullulans in the ITS sequence, has the morphological characteristic of forming dark black colonies, and contains polysaccharides such as xylan, arabinoxylan, amylose, and xyloglucan. There is also a significant difference compared to the existing Aureobasidium pullulans in that it has metabolic characteristics of decomposing xylan and producing disaccharides as the main product.

The strain of the present invention can exhibit the above-mentioned polysaccharide decomposition activity and the activity of producing disaccharide from xylan outside the cell. This was confirmed through the results of experiments on the decomposition activity of polysaccharides (including xylan) using bacterial removal culture media, suggesting that enzymes related to this activity, such as xylanase, can be secreted to the outside of the cell.

The strain of the present invention can be cultured according to a conventional culture method for culturing fungi, especially fungi of the Aureobasidium genus, especially fungi of the Aureobasidium pullulans species. For example, for solid culture, PDA (potato dextrose agar) medium can be used, and for liquid culture, PDB (potato dextrose broth) medium can be used, and the culture can be done under temperature conditions of 20 to 40°C.

Based on the characteristics of the strain of the present invention as described above, the present invention provides a method of decomposing polysaccharides comprising the step of contacting a culture of the strain of the present invention or a purified product of the culture with a polysaccharide.

In the present invention, a culture of the present strain can be obtained by culturing the present strain according to the culture method described above. Preferably, the culture of the strain of the present invention is obtained by culturing in PDB medium. More preferably, the culture of the strain of the present invention is obtained by culturing in PDB medium at 28 to 32 ℃ for 2 to 4 days, and more preferably, it is obtained by culturing at 29 to 31 ℃ for 60 to 84 hours. will be. According to this, a culture with higher polysaccharide decomposition activity can be obtained. The culture of the strain of the present invention is preferably obtained by removing the bacterial cells from a culture grown in a liquid medium. This can be achieved using conventional methods for removing bacterial cells from the culture, such as centrifugation and filtration.

In the present invention, the purified product of the culture is obtained through the purification process of the culture of the strain of the present invention, and is obtained from a culture of microorganisms, especially fungi, especially fungi of the genus Aureobasidium, especially fungi of the species Aureobasidium pullulans. It may be obtained through a general purification process used to purify externally secreted proteins, especially enzymes, especially glycolytic enzymes. For example, it may be obtained by filtering the culture medium from which the bacterial cells have been removed with a 10KDa cut-off filter.

In the polysaccharide decomposition method of the present invention, the step of contacting the polysaccharide is preferably a step of contacting the culture of the strain of the present invention or a purified product of the culture with xylan, arabinoxylan, amylose or xyloglucan, more preferably This is the step of contacting xylan.

Contact between the culture of the strain of the present invention or a purified product of the culture and the polysaccharide can be carried out by adding polysaccharide to the culture or purified product, or can be done in a buffer solution. For example, it can also be performed by adding the culture or purification and polysaccharide to a buffer solution. The contact may be carried out, for example, at a temperature of 30 to 40°C, and is preferably carried out at 35 to 39°C.

Based on the above characteristics of the strain of the present invention, the present invention also provides a composition for decomposing polysaccharides comprising a culture of the strain of the present invention or a purified product of the culture.

Details regarding culture and purification in the composition for decomposing polysaccharides of the present invention can be referred to the information described above.

The composition for decomposing polysaccharides of the present invention may further include other ingredients in addition to the culture or purification. For example, ingredients that do not substantially inhibit the decomposition activity of polysaccharides by the culture or purification may further include buffers, diluents, excipients, etc. for the stable action of glycolytic enzymes.

The composition for decomposing polysaccharides of the present invention may include, for example, 0.001 to 100% of the culture or purified product based on the total weight of the composition.

Based on the above characteristics of the strain of the present invention, the present invention also provides a method for producing a disaccharide comprising the step of contacting a culture of the strain of the present invention or a purified product of the culture with xylan. This is based on experimental results showing that the strain of the present invention can decompose xylan and produce disaccharide as the main product, and that this activity can be exerted in a culture medium from which the bacteria have been removed.

Details regarding culture and purification in the disaccharide production method of the present invention can be referred to the information described above.

For matters regarding contact between a culture of the strain of the present invention or a purified product of the culture and xylan, reference may be made to the description of contact between a culture of the strain of the present invention or a purified product of the culture and polysaccharide.

The disaccharide production method of the present invention may further include the step of purifying the disaccharide produced after contacting it with xylan. At this time, for purification matters, refer to the usual method of purifying a specific reaction product from the reaction mixture after the enzymatic reaction of polysaccharide.

Based on the above characteristics of the strain of the present invention, the present invention also provides a composition for producing disaccharides comprising a culture of the strain of the present invention or a purified product of the culture.

Details regarding culture and purification in the composition for producing disaccharides of the present invention can be referred to the information described above.

The composition for producing disaccharides of the present invention may further include other ingredients in addition to the culture or purification. For example, ingredients that do not substantially inhibit the decomposition activity of xylan by the culture or purification may further include buffers, diluents, excipients, etc. for stable action of the glycolytic enzyme.

The composition for producing a disaccharide of the present invention may include, for example, 0.001 to 100% of the culture or purified product based on the total weight of the composition.

Hereinafter, the present invention will be described in more detail through examples. Since these examples are merely for illustrating the present invention, the scope of the present invention is not to be construed as limited by these examples.

[Example]

1. Method

1-1. Strain isolation

One rose flower collected from around Iui-dong, Yeongtong-gu, Gyeonggi-do was sterilized by immersing it in 200 ml of 80% ethanol for 10 seconds and washed four times in 500 ml of sterilized water. 50 ㎖ of sterilized water was added to the sample from which water was removed, and the sample was thoroughly crushed in a mortar. 200 ㎕ of the supernatant was spread on MRS solid medium and cultured at 25°C for 2 days to obtain the resulting colonies.

A total of 11 colonies were obtained, classified as fungi based on their morphology, transferred to PDA solid medium containing 0.2% AZCL (Azurine-crosslinked)-xylan, and cultured at 25°C for 5 days. The decomposition activity was confirmed, and one strain with strong activity was finally selected.

The final selected strain was named RG4.

1-2. Strain identification

To identify RG4, the ITS region was sequenced using primers ITS1F (5'-CTTGGTCATTTAGAGGAAGTAA-3') and ITS4 (5'-TCCTCCGCTTATTGATATGC-3') and NL1 (5'-GCATATCAATAAGCGGAGGAAAAG-3'). And the nucleotide sequence of the D1D2 region was analyzed using primers NL4 (5'-GGTCCGTGTTTCAAGACGG-3').

Based on the nucleotide sequences of the ITS region and the D1D2 region, homology analysis of the nucleotide sequence was performed from the GenBank database information using the BlastN program of the National Center for Biotechnology Information (NCBI). For phylogenetic tree analysis, the ITS-D1D2 sequences of strains showing homology were obtained, and a NJ (Neighbor-Joining) phylogenetic tree was created using the Mega X program to analyze phylogenetic relationships.

1-3. Analysis of morphological and physiological characteristics

RG4 was cultured in PDA medium to observe the changing morphology of colonies as they grew on the medium, and hyphae were observed using an Axio Imager Z2 microscope (Zeiss) on the 7th day of culture.

Physiological characteristics were investigated using API 20NE, API 20E, and API ZYM kits (Biomerieux, France) according to the presented method. The bacterial sample was obtained by culturing RG4 in PDA medium for 2 days.

1-4. Polysaccharide degradation activity analysis

To determine the polysaccharide decomposition activity of RG4, RG4 was treated with PDA containing 0.2% AZCL-xylan, -amylose, -arabinoxylan, -xyloglucan, and -cellulose, respectively. After smearing the medium and culturing it at 25°C for 48 hours, it was observed that the AZCL substrate around the colonies was decomposed and turned blue.

RG4 was inoculated into 200 ml of PDB liquid medium and cultured at 30°C for 10 days. 5 ml of the culture was sampled every 3 days, and centrifuged at 15,000 rpm for 30 minutes to remove the bacterial cells. Only the supernatant was taken. The prepared supernatant was filtered through a 0.22㎛ syringe filter and used as an enzyme solution, and xylanase activity was measured using the DNS (dinitrosalicylic acid) method [Miller GL. 1959. Use of dinitrosalicylic acid reagent for determination of reducing sugar. Anal. Chem. 31: 426-428.], and the amount of reducing sugar liberated by the enzyme reaction was measured at an absorbance of 540 nm.

1-5. TLC analysis

To analyze the hydrolysis products produced by RG4 by decomposing xylan, samples from the 3rd day, which showed the highest activity as a result of measuring enzyme activity by bacterial growth period, were filtered through an Amicon ultracentrifugal filter (10KDa cut-off, Millipore, USA) was used to prepare a 10-fold concentrated crude enzyme solution.

To analyze xylan degradation products, 100㎕ of crude enzyme solution was added to 1X phosphate buffered saline (PBS) containing 0.4% xylan and reacted at 37°C for 24 hours. During the enzyme reaction, the enzyme reaction solution was sampled at 12-hour intervals and 20 μl was added to a silica gel 60 plate (Merck). At this time, xylobiose, xylotetraose, and xylohexaose (Megazyme, Ireland) were used as standards. A solution of n-butanol: acetic acid: distilled water (2:1:2) was used as a developing solvent, and a solution of ethanol: sulfuric acid (9:1) was used as a color developing reagent. After development, the color development reagent was evenly sprinkled and confirmed by heating at 120°C.

2. Results

2-1. Isolation and identification of RG4

RG4 was finally selected as the strain with the best xylan decomposition activity among the strains forming dark black colonies selected from rose flower samples.

When RG4 is cultured in PDA medium, it is cream-colored at first, grows sticky, and turns black overall over time. Under the microscope, hyphae and oval shapes (similar to the shape of general yeast) coexisted, and regular spores and black chlamydospores could also be observed (Figure 2).

By analyzing the ITS-D1D2 sequence of RG4, it was classified as a species of the genus Aureobasidium . As a result of NCBI Blast homology analysis, RG4 showed high homology with Aureobasidium genus strains, especially the highest homology with Aureobasidium pullulans CBS 584.75 (ITS: 99.14%, D1D2: 100%) (Table 1). However, when Aureobasidium pullulans CBS 584.75 is cultured in PDA medium, it does not secrete black pigment and does not show the color of the cells, whereas RG4 of the present invention shows a dark black color toward the center of the colony. There was a clear difference in enemy characteristics. In addition, Aureobasidium subglaciale ( A. subglaciale ), Aureobasidium namibiae ( A. namibiae ), and Aureobasidium, which were classified as Aureobasidium pullulans but were recently reclassified as other species. It also showed many differences in morphological characteristics from A. melanogenum (Redefinition of Aureobasidium pullulans and its varieties 2008. Studies in Mycology. 61: 21-38). Based on these homology search results, RG4 was identified as Aureobasidium pullulans, but as a result of observation of morphological characteristics, it was found to be different from the previously known standard strain, Aureobasidium pullulans CBS 584.75, and its varieties. It was found to be a new strain. Finally, the strain RG4 discovered in the present invention was named Aureobasidium pullulans RG4.

As a result of enzyme activity analysis using the API ZYM kit, Aureobasidium pullulans RG4 showed positive reactions in all activities except α-mannosidase activity (Table 2). API 20E kit analysis results showed enzyme activities such as beta-galactosidase, urease, tryptophan deaminase, and gelatinase, and carbon sources such as D-glucose, L-rhamnose, D-sucrose, D-mellibiose, and amygdalin. The fermentation reaction was observed to be positive (Table 3). As a result of the API 20NE kit analysis, D-glucose fermentation reaction was positive, esculin decomposition, gelatinase, beta-galactosidase, etc. enzyme activities, D-glucose, L-arabinose, D-mannose, and D-mannitol. , was observed positively in the use of carbon sources such as N-acetyl-glucosamine, D-maltose, and malic acid (Table 4).

ITS-D1D2 nucleotide sequence homology search results of RG4 scientific name isolate ITS sequence registration number sameness(%) D1D2 sequence registration number sameness(%) Aureobasidium pullulans CBS 584.75 ^T NR_144909 99.14 NG_055734 100 Aureobasidium subglaciale ^CBS123387T NR_147323 98.05 NG_058546 96.96 Aureobasidium namibiae CBS 147.97 ^T NR_147362 98.27 NG_056961 97.53 Aureobasidium melanogenum CBS ^105.22T NR_159598 97.55 NG_056960 97.34 Aureobasidium lini CBS ^125.21T NR_147320 98.16 NG_056962 99.43 Aureobasidium mangrovei IBRC-M- ^30265T KY089085 92.87 KY089084 96.58 Aureobasidium iranianum CCTU ^268T NR_137598 95.74 NG_057049 96.77 Aureobasidium thailandense NRRL ^58539T NR_147337 92.70 JX462674 95.83 Aureobasidium leucospermi ^CBS130593T NR_156246 98.20 NG_058566 97.15 Selenophoma linicola CBS 468.48 ^T NR_160075 90.29 NG_057801 94.89

API ZYM kit analysis results of Aureobasidium pullulans RG4 enzyme reaction Alkaline phosphatase + Esterase(C4) + Esterase Lipase(C8) + Lipase(C14) + Leucine arylamidase + Valine arylamidase + Cystine arylamidase + Trypsin +- α-chymotrypsin +- Acid phosphatase + Naphtol-AS-BI-phosphohydrolase + α-galactosidase + β-galactosidase + β-glucuronidase + α-glucosidase + β-glucosidase + N-acetyl-D-β-glucosaminidase + α-mannosidase - α-fucosidase +

Symbol: +, positive; +-, weakly positive; -, voice.

API 20E kit analysis results of Aureobasidium pullulans RG4 Enzyme/Substrate reaction enzyme β-galactosidase + Arginine dihydrolase - Lysine decarboxylase - Ornithine decarboxylase - Citrate utilization - H₂S production - Urease + Tryptophane deaminase + indole production - acetoin production (Vogesproskauer) - Gelatinase + fermentation D-glucose + D-mannitol - inositol - D-sorbitol - L-rhamnose +- D-sucrose + D-melibiose +- amygdalin + L-arabinose -

Symbol: +, positive; +-, weakly positive; -, voice.

API 20NE kit analysis results of Aureobasidium pullulans RG4 Enzyme/Substrate reaction enzyme Nitrates to nitrites - Indole production - Arginine dihydrolase -+ Urease - Esculin hydrolysis - Gelatin hydrolysis + Galactosidase + fermentation Glucose +- Fairytale D-glucose + L-arabinose + D-mannose + D-mannitol + N-acetyl-glucosamine +- D-maltose + potassium gluconate - capric acid - adipic acid - malic acid + trisodium citrate - phenylacetic acid -

Symbol: +, positive; +-, weakly positive; -, voice.

2-2. Polysaccharide decomposition activity

Aureobasidium pullulans RG4 decomposes insoluble AZCL-amylose, AZCL-arabinoxylan, AZCL-xylan, and AZCL-xyloglucan around the cell, causing a blue color in the medium, and secretes decomposing enzymes to the outside of the cell. I could see that Among the four enzyme activities, xylan decomposition activity was observed to be the strongest, followed by arabinoxylan decomposition activity, followed by amylose decomposition activity and xyloglucan decomposition activity. On the other hand, no decomposition activity was observed for AZCL-cellulose (Figure 3).

When measuring xylanase activity according to liquid culture of Aureobasidium pullulans RG4 in PDB medium, the highest enzyme activity was seen in the sample on the 3rd day, but rapidly decreased from the 5th day (Figure 4).

When the enzyme reaction product using xylan as a substrate was analyzed by TLC using the crude enzyme solution prepared from the sample on the 3rd day of liquid culture of Aureobasidium pullulans RG4, the xylanase secreted by Aureobasidium pullulans RG4 was not present. It was found that beechwood xylan was decomposed to produce dimer (DP2) as the main decomposition product (FIG. 5). It was confirmed that oligosaccharides with a structure longer than the hexamer (DP6) were produced, including the dimer, which is the final decomposition product, and this TLC pattern is consistent with the hydrolysis characteristics of a general endo-type xylanase. Therefore, it is believed that if the enzyme of Aureobasidium pullulans RG4 is used, it will be possible to produce xylooligosaccharide with dimers as the main product from xylan.

Agricultural Biotechnology Research Institute KACC93361P 20210615

<110> National Institute of Biological Resources <120> Novel strain Aureobasidium pullulans RG4 having xylan degrading activity <130> ALP21011 <160> 2 <170> KoPatentIn 3.0 <210> 1 <211> 550 <212> DNA <213> Unknown <220> <223> Aureobasidium pullulans RG4 <400> 1 ggataaaagg actcacgccc gacctccaac cctttgttgt taaaactacc ttgttgcttt 60 ggcgggaccg ctcggttccg agccgctggg gattcgtccc aggcgagcgc ccgccagagt 120 taaaccaaac tcttgttat taaccggtcg tctgagtaaa aattttgaat aaatcaaaac 180 tttcaacaac ggatctcttg gttctcgcat cgatgaagaa cgcagcgaaa tgcgataagt 240 aatgtgaatt gcagaattca gtgaatcatc gaatctttga acgcacattg cgccccttgg 300 tattccgagg ggcatgcctg ttcgagcgtc attacaccac tcaagctatg cttggtattg 360 ggcgtcgtcc ttagttgggc gcgccttaaa gacctcggcg aggccactcc ggctttaggc 420 gtagtagaat ttattcgaac gtctgtcaaa ggagaggaac tctgccgact gaaaccttta 480 tttttctagg ttgacctcgg atcaggtagg gatacccgct gaacttaagc atatcataan 540 ccggaggaaa 550 <210> 2 <211> 526 <212> DNA <213> Unknown <220> <223> Aureobasidium pullulans RG4 <400> 2 aacagggatt gccctagtaa cggcgagtga agcggcaaca gctcaaattt gaaagctagc 60 cttcgggttc gcattgtaat ttgtagagga tgatttgggg aagccgcctg tctaagttcc 120 ttggaacagg acgtcataga gggtgagaat cccgtatgtg acaggaaatg gcaccctatg 180 taaatctcct tcgacgagtc gagttgtttg ggaatgcagc tctaaatggg aggtaaattt 240 cttctaaagc taaatattgg cgagagaccg atagcgcaca agtagagtga tcgaaagatg 300 aaaagcactt tggaaagaga gttaaaaagc acgtgaaatt gttgaaaggg aagcgcttgc 360 aatcagactt gtttaaactg ttcggccggt cttctgaccg gtttactcag tttggacagg 420 ccagcatcag tttcggcggc cggataaagg ctctgggaat gtggcctcca cttcggtgga 480 ggtgttatag cccaggggtgt aatacggcca gccgggactg aggtcc 526

Claims (7)

Aureobasidium pullulans RG4 strain deposited with accession number KACC 93361P. A polysaccharide decomposition method comprising the step of contacting a culture of the strain of claim 1 or a purified product of the culture with a polysaccharide,
A method of decomposing a polysaccharide, wherein the polysaccharide is xylan, arabinoxylan, amylose or xyloglucan. delete A composition for decomposing polysaccharides comprising a culture of the strain of claim 1 or a purified product of the culture,
A composition for decomposing a polysaccharide, wherein the polysaccharide is xylan, arabinoxylan, amylose, or xyloglucan. delete A disaccharide production method comprising the step of contacting a culture of the strain of claim 1 or a purified product of the culture with xylan. A composition for producing a disaccharide comprising a culture of the strain of claim 1 or a purified product of the culture.