Application of xylanase

Abstract

The invention discloses application of xylanase, and belongs to the technical field of bioengineering. According to preferred codons of bacillus subtilis, a wild type xylanase (GH11 family) coding gene which is secreted by Cellulostridium morganii and is not represented is designed, and a genetic engineering technology is utilized, so that the wild type xylanase is subjected to recombinant expression in the bacillus subtilis. Through verification, the separated xylanase still has xylan hydrolysis activity under the conditions of high temperature (close to 100 DEG C) and strong alkali (pH 9-10.5), and shows good heat resistance and alkali resistance; and meanwhile, more than 60% of enzyme activity can still be maintained in a high-temperature (100 DEG C) environment within 1 hour, and the strain has relatively high thermal stability and has a good application prospect in extreme environments such as industrial production and the like.

Description

Application of a xylanase Technical Field

[0001] This invention belongs to the field of bioengineering technology, specifically relating to the application of a xylanase. Background Technology

[0002] Lignocellulose, as one of the most abundant renewable biological resources, is mainly composed of three components: cellulose, hemicellulose, and lignin. Hemicellulose is a heteropolysaccharide composed of monosaccharides such as glucose, mannose, galactose, xylose, and arabinose. Xylose is linked by glycosidic bonds to form xylan, which, as the main component of hemicellulose, can account for up to 35% of lignocellulosic biomass (such as straw and wood). In addition to its abundant source, xylan also possesses good biodegradability and is considered the second most abundant renewable polysaccharide resource after cellulose.

[0003] Xylanases belong to the glycoside hydrolase (GH) family, with the most crucial GH11 family xylanase being endo-β-1,4-D-xylanase (EC 3.2.1.8). This enzyme degrades the β-1,4-xylosidic bonds within the xylan backbone, hydrolyzing xylan to produce xylooligosaccharides and xylose. Due to their high catalytic efficiency and environmental friendliness, xylanase-based enzymatic methods have gradually replaced traditional chemical catalytic methods in industrial production fields such as textiles, food, feed, papermaking, and lignocellulose biomass conversion. However, actual industrial production may encounter extreme reaction conditions such as high temperatures and strong alkalis, which places higher demands on the application of most xylanases, whose optimal reaction temperatures are between 30 and 60°C and pH values ​​between 3.0 and 6.0, in extreme environments. Traditional enzyme molecule modification technologies (including heat resistance and acid / alkali resistance modification) are technically difficult and costly, which reduces the economic efficiency of industrial production to some extent.

[0004] In summary, exploring xylanases that are heat-resistant and have a wider range of applications is of great significance for the industrial production of more extremophile enzymes. Summary of the Invention

[0005] To address the issue that most existing xylanases exhibit poor stability under extreme conditions such as high temperatures and strong alkalis, and that modifying them using enzyme molecular modification technology is costly, significantly limiting their application in extreme environments such as industrial production, this invention provides a heat- and alkali-resistant xylanase suitable for use in extreme environments such as high temperatures (near 100°C) and strong alkalis (pH 9-1.0.5). Based on the codons preferred by Bacillus subtilis, the coding gene of an uncharacterized wild-type xylanase (GH11 family) derived from *Caldicellulosiruptor morganii* was designed. Using genetic engineering technology, this gene was recombinantly expressed in *Bacillus subtilis*. The resulting xylanase exhibits excellent xylan catalytic activity, effectively hydrolyzing xylan at temperatures near 100°C, and high thermal stability (retaining over 60% of its activity within 1 hour). Furthermore, it possesses good alkali resistance, making it suitable for extreme environments such as industrial production.

[0006] To achieve the above-mentioned objectives, the technical solution adopted by this invention is as follows:

[0007] This invention provides an application of xylanase, which includes: reacting xylanase with xylan or raw materials containing xylan under extreme conditions to achieve xylan hydrolysis; the extreme conditions include: a temperature of 60-100℃ and / or a pH of 6-10.5; the amino acid sequence of the xylanase is shown in SEQ ID NO.1; it should be noted that this invention uses heterologous expression of uncharacterized wild-type xylanase (GH11 family) secreted by *Caldicellulosiruptor morganii*. The isolated xylanase has been verified to have heat and alkali resistance, and still exhibits high enzymatic activity for xylan hydrolysis under conditions as high as 60-100℃ and pH 6-10.5.

[0008] Furthermore, the aforementioned extreme environments include temperatures of 70–100°C and / or pH values ​​of 7–10.5.

[0009] Furthermore, the aforementioned extreme environments include temperatures of 80–100°C and / or pH values ​​of 8–10.5.

[0010] Furthermore, the aforementioned extreme environments include temperatures of 90–100°C and / or pH values ​​of 9–10.5.

[0011] Furthermore, the aforementioned raw materials containing xylan include any one or more of the following: grain raw materials, agricultural waste, forestry raw materials, and textile raw materials.

[0012] Furthermore, the optimal temperature for the activity of the above-mentioned xylanase is 80~90℃, and the optimal pH is 6~7.

[0013] The present invention also provides a method for preparing xylooligosaccharides and / or xylose under extreme conditions, the method comprising: adding xylanase with an amino acid sequence as shown in SEQ ID NO.1 to xylan or a raw material containing xylan components, and carrying out a contact reaction at a temperature of 60~100℃ and / or pH 6~10.5.

[0014] Furthermore, the above-mentioned xylanase is added to xylan or raw materials containing xylan components, and a contact reaction is carried out at a temperature of 70~100℃ and / or pH 7~10.5.

[0015] Furthermore, the above-mentioned xylanase is added to xylan or raw materials containing xylan components, and a contact reaction is carried out at a temperature of 80~100℃ and / or pH 8~10.5.

[0016] Furthermore, the above-mentioned xylanase is added to xylan or raw materials containing xylan components, and a contact reaction is carried out at a temperature of 90~100℃ and / or pH 9~10.5.

[0017] Furthermore, the aforementioned raw materials containing xylan include any one or more of the following: grain raw materials, agricultural waste, forestry raw materials, and textile raw materials.

[0018] Furthermore, the optimal temperature for the activity of the above-mentioned xylanase is 80~90℃, and the optimal pH is 6~7.

[0019] The present invention also provides an enzyme composition suitable for extreme environments, the composition comprising xylanase with the amino acid sequence shown in SEQ ID NO.1, which has enzymatic activity for catalyzing the hydrolysis of xylan under conditions of temperature 60-100°C and / or pH 6-10.5.

[0020] Furthermore, the above-mentioned xylanase exhibits enzymatic activity catalyzing the hydrolysis of xylan under conditions of temperature 70~100℃ and / or pH 7~10.5.

[0021] Furthermore, the above-mentioned xylanase exhibits enzymatic activity catalyzing the hydrolysis of xylan under conditions of temperature 80-100℃ and / or pH 8-10.5.

[0022] Furthermore, the above-mentioned xylanase exhibits enzymatic activity catalyzing the hydrolysis of xylan under conditions of temperature 90-100℃ and / or pH 9-10.5.

[0023] Compared with the prior art, the beneficial effects of this invention are as follows: (1) The application of the xylanase provided by this invention involves designing the encoding gene of an uncharacterized wild-type xylanase (GH11 family) secreted by *Caldicellulosiruptor morganii*, and using genetic engineering technology to recombinantly express it in *Bacillus subtilis*. The isolated xylanase has been proven to be able to efficiently hydrolyze xylan under high temperature (close to 100℃) and strong alkaline (pH 9~10.5) conditions, and has good heat and alkali resistance. At the same time, the xylanase also exhibits good thermal stability, and can still maintain more than 60% of its enzyme activity within 1 hour at high temperature (100℃), showing good application prospects in extreme environments such as industrial production.

[0024] (2) The application of xylanase provided by the present invention is different from the traditional method of modifying xylanase by enzyme molecule modification technology to improve its thermal stability and alkali resistance. The present invention can obtain xylanase with high purity (greater than 95%), good thermal stability and alkali resistance by heterologous expression. The whole operation process is simple, the technology is relatively mature, and it has good economic benefits. It is suitable for use in extreme environments such as large-scale production and industrial production. Attached Figure Description

[0025] Figure 1 shows the protein electrophoresis results of the crude xylanase solution and the purified xylanase solution in this invention. From left to right, the lanes correspond to the following samples: Marker, crude xylanase solution, and purified xylanase solution. The sizes of the Marker bands from top to bottom are: 140 KD, 115 KD, 80 KD, 65 KD, 50 KD, 40 KD, 30 KD, 25 KD, and 15 KD. The purified xylanase is approximately 26 KD in size.

[0026] Figure 2 is a standard curve of xylose in this invention.

[0027] Figure 3 shows the relative enzyme activities of xylanase at different temperatures in this invention.

[0028] Figure 4 shows the relative enzyme activities of xylanase at different pH values ​​in this invention.

[0029] Figure 5 shows the results of the xylanase thermostability test in this invention. Detailed Implementation

[0030] The present invention will be further described below with reference to specific embodiments.

[0031] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains; the term “and / or” as used herein includes any and all combinations of one or more of the associated listed items.

[0032] Unless otherwise specified in the examples, the procedures should be performed under standard conditions or conditions recommended by the manufacturer. Reagents or instruments whose manufacturers are not specified are all commercially available products.

[0033] As used herein, the term “about” is used to provide for the flexibility and imprecision associated with a given term, measure, or value. Those skilled in the art can readily determine the degree of flexibility for a particular variable.

[0034] As used herein, the term “at least one of…” is intended to be synonymous with “one or more of…”. For example, “at least one of A, B, and C” explicitly includes only A, only B, only C, and combinations thereof.

[0035] Concentration, amount, and other numerical data may be presented in range format herein. It should be understood that such range format is used solely for convenience and brevity and should be flexibly interpreted to include not only the values ​​explicitly stated as the limits of the range, but also all individual values ​​or subranges encompassed within the range, as if each value and subrange were explicitly stated. For example, a range of values ​​from about 1 to about 4.5 should be interpreted to include not only the explicitly stated limits of 1 to 4.5, but also individual numbers (such as 2, 3, 4) and subranges (such as 1 to 3, 2 to 4, etc.). The same principle applies to ranges that describe only a single value, such as "less than about 4.5," which should be interpreted to include all the values ​​and ranges described above. Furthermore, this interpretation should apply regardless of the breadth of the range or characteristic described.

[0036] The reagents and culture media used in this invention were prepared as follows: 0.1 M citrate buffer (pH 6.5): 21.0 g of citric acid monohydrate was dissolved in 950 mL of deionized water, and 1.0 M NaOH was added to adjust the pH to 6.5, followed by a final volume adjustment to 1000 mL; LB liquid medium: 10 g / L peptone; 5 g / L yeast extract; 10 g / L NaCl; LB solid medium: 2% agarose was added to the above LB liquid medium formulation; xylan: purchased from Sigma, catalog number 9014-63-5; xylose: purchased from Heinz, catalog number 100-X99999KDW1; 3,5-dinitrosalicylic acid: purchased from Yuanye Biotechnology, catalog number R27125; cellulase: purchased from Megazyme, catalog number E-CELAN.

[0037] The instrument used in this embodiment of the invention: Ultraviolet spectrophotometer: Tianmei UV2500 model.

[0038] Example 1 This example provides screening for wild-type xylanase.

[0039] The wild-type xylanase was screened from an uncharacterized glycoside hydrolase family 11 (GH11) secreted by *Caldicellulosiruptormorganii*, and its amino acid sequence is shown in SEQ ID NO:1 (NCBI Reference Sequence: WP_052670803.1). "Wild-type" refers to the form found in nature, which can be isolated from nature and has not undergone artificial modification. Based on the codon preference of *Bacillus subtilis*, the encoding gene for the above-mentioned xylanase was designed, and the nucleotide sequence shown in SEQ ID NO:2 was obtained through codon optimization.

[0040]

[0041] Example 2 This example provides the preparation of xylanase.

[0042] To characterize the function of the wild-type xylanase in Example 1, xylanase was prepared and purified by heterologous expression in this example.

[0043] The specific steps are as follows: (1) Synthesize the target gene The optimized nucleotide sequence (SEQ ID NO.2) of the codon in Example 1 is synthesized using whole gene synthesis technology. In order to realize the construction of the recombinant expression vector, BamHI and SalI restriction endonuclease sites are designed at both ends of the target gene. The above work was entrusted to Anshengda Biotechnology Co., Ltd.

[0044] (2) Construction of recombinant expression vector The vector used in this embodiment is plasmid pWB980. Specifically, the target gene and pWB980 synthesized above were double-digested with BamHI and SalI restriction endonucleases, respectively, and then identified by agarose gel electrophoresis. The digested target gene fragment and pWB980 were recovered by gel excision and ligated with T4 DNA ligase. The ligation product was transformed into competent DH5α cells. Positive clones were screened using 50 μg / mg kanamycin, indicating that the recombinant expression vector was successfully constructed and named pXyn1.

[0045] (3) Construction of genetically engineered bacteria: The recombinant expression vector pXyn1 was transformed into Bacillus subtilis 168 by electroporation. The specific operation was as follows: Bacillus subtilis 168 competent cells stored at -80℃ were taken out and placed on ice. 500 ng of recombinant expression vector was mixed with 80 μL of competent cells and incubated on ice for 2 min. Then, it was transferred to a pre-cooled electroporation cuvette (1 mm). The electroporation instrument conditions were set as follows: 2.0 kV, 1 mm, 1 electroporation. After electroporation, 1 mL of LB liquid medium (containing 1 M sorbitol, 50% by volume) was immediately added to the electroporation cuvette. After recovery at 37℃ and 200 rpm for 2 h, it was spread on LB solid medium containing 50 μg / mL kanamycin and incubated upside down at 37℃ overnight. Single clones were picked from the plate and inoculated into LB liquid medium for expansion culture. The recombinant engineered bacterial culture was stored at -80℃ with 25% final concentration of glycerol for later use. Some of the culture was sent to Anshengda Biotechnology Co., Ltd. for testing. The sequence analysis confirmed that the culture was correct, and the xylanase recombinant engineered bacterial culture was successfully constructed. In this invention, it is named BS(pXyn1).

[0046] (4) Expanding the culture: The above-mentioned recombinant engineered bacteria BS (pXyn1) was inoculated into LB liquid medium (containing 100 μg / mL kanamycin) and cultured overnight at 37℃ and 220 rpm. The culture was then transferred to 50 mL of fresh LB liquid medium (250 mL shake flask) at a ratio of 1:100 and grown at 37℃ and 220 rpm for 24 h. The fermentation broth was then centrifuged at 12000 rpm and 4℃ for 10 min, and the supernatant, i.e., crude enzyme solution, was collected and stored at -20℃ for later use.

[0047] (5) Isolation and purification of xylanase: The above crude enzyme solution was isolated and purified by ion exchange method. The ion exchange column used was a HiTrap Q FF (5 mL) strong anion exchange column; the buffer A was prepared as follows: 2.42 g Tris (tris(hydroxymethyl)aminomethane) was weighed and dissolved in 1 L of deionized water, and the pH was adjusted to 8.0 with hydrochloric acid to obtain a 20 mM Tris-HCl buffer; the buffer B was prepared as follows: 2.42 g Tris was weighed and dissolved in 1 L of deionized water, and 58.44 g NaCl (sodium chloride) was added, and the pH was adjusted to 8.5 with hydrochloric acid to obtain a 20 mM Tris-HCl, 1 M NaCl buffer.

[0048] The specific purification process is as follows: First, wash with buffer B at a flow rate of 2.5 mL / min for 3 column volumes (CV); then equilibrate with buffer A at the same flow rate for 10 CV; then load the sample using a loading loop at the same flow rate of 2.5 mL / min; next, wash with buffer A at the same flow rate for 8 CV; finally, perform linear gradient elution with buffer B, 0~40% B, 10 CV, while maintaining the flow rate at 2.5 mL / min, to obtain purified xylanase enzyme solution.

[0049] (6) Xylanase detection: The crude xylanase solution and the purified xylanase solution were subjected to polyacrylamide gel electrophoresis. The sample loading volume for each lane was 5 μL, and the electrophoresis conditions were 120 V constant voltage for 40 min.

[0050] The results are shown in Figure 1. From left to right, the lanes correspond to the marker, crude xylanase solution, and purified xylanase solution, respectively. It can be seen that the molecular weight of the purified xylanase is approximately 26 KD, consistent with the molecular weight estimated from its amino acid sequence, indicating successful preparation of xylanase. The integrated optical density was calculated using Gel-Pro software, showing that the purity of the purified xylanase was greater than 95%.

[0051] Example 3 This example provides a method for determining the enzyme activity of xylanase.

[0052] The 3,5-dinitrosalicylic acid (DNS) colorimetric method was used to reflect the enzyme activity of xylanase by calculating the amount of reducing sugar (calculated as xylose) generated during the reaction between the xylan substrate and the enzyme solution. The enzyme activity unit is defined as: the amount of enzyme required to produce 1 μmol of reducing sugar (xylose) per minute by hydrolyzing 1 mL of enzyme solution under certain conditions is 1 enzyme activity unit, i.e., U / mL. The specific determination process is as follows: (1) The principle on which the xylose standard curve is based: 3,5-dinitrosalicylic acid (DNS) reacts with reducing sugar (xylose) under alkaline conditions to generate 3-amino-5-nitrosalicylic acid. This product is brownish-red under boiling conditions and has maximum absorption at a wavelength of 540 nm. Moreover, within a certain concentration range, the color intensity is proportional to the reducing sugar content. Therefore, the xylose content is determined by colorimetry.

[0053] Specifically, in this embodiment, xylose solutions were prepared according to the xylose concentration gradient set in Table 1. DNS was then added to the solution, and after mixing, the mixture was reacted for 30 min. The solution was then boiled in boiling water for 10 min, cooled to room temperature with cold water, and the absorbance of the reaction solution at 540 nm was measured using a UV spectrophotometer. A xylose standard curve was plotted with xylose concentration (mg / mL) on the x-axis and absorbance (A) on the y-axis. The results are shown in Figure 2, with a correlation coefficient R... 2 A value greater than 0.99 indicates a good linear fit.

[0054] Table 1. Composition of the reaction system used in plotting the xylose standard curve

[0055] (2) The final system for each group of enzyme activity determination experiments was set to 500 μL, as follows: Enzyme reaction group: including 50 μL of 0.1 M pH 6.0 citrate buffer, 250 μL of xylan (1% W / V), and 200 μL of xylanase purified enzyme solution; Enzyme blank group: including 50 μL of 0.1 M pH 6.0 citrate buffer, 250 μL of xylan (1% W / V), and 200 μL of inactivated xylanase purified enzyme solution; The purpose of setting up this group is to deduct the background value of the system caused by non-enzymatic reactions such as spontaneous decomposition of substrate and oxidation of reagents in the system, and to correct the actual reaction value; Zeroing system group: 500 μL of 0.1 M pH 6.0 citrate buffer. The purpose of setting up this group is to zero the UV spectrophotometer before testing the sample.

[0056] The enzyme reaction group and the enzyme blank group were subjected to a water bath reaction under the same conditions for 30 min. Then, 200 μL of DNS was immediately added, and the reaction was carried out in a metal bath at 100°C for 10 min. After cooling in a 4°C refrigerator for 5 min, the absorbance at 540 nm was measured using a UV spectrophotometer. Then, based on the linear regression equation fitted to the xylose standard curve, the xylose content in the enzyme reaction solution was calculated using the following formula:

[0057] In the formula: C 木糖 Xylose content (μmol / L) in the enzyme reaction solution; A 反应液 A represents the absorbance of the enzyme reaction solution. 空白 ρ represents the absorbance of the enzyme blank group; a is the slope of the linear regression equation, b is the intercept of the linear regression equation, and M is the value of the enzyme blank group. 木糖 The molar mass of xylose is 150.13 g / mol; the enzyme activity of xylanase is then calculated using the following formula:

[0058] Where: enzyme activity (U / mL); C 木糖 Xylose content in the enzyme reaction solution (μmol / L); V 总 V is the total volume of the enzyme reaction solution (L); 酶 T represents the volume of enzyme solution added (mL); T represents the reaction time (min); and N represents the dilution factor of the enzyme solution.

[0059] Example 4 This example uses the enzyme activity detection method in Example 3 to investigate the catalytic characteristics of xylanase.

[0060] (a) The optimal temperature is the same as the reaction system in Example 3. The difference in this example is that the pH of the citrate buffer in each system is adjusted to 6.5, while other components remain unchanged. The enzyme solution used is xylanase purified enzyme solution diluted 5000 times. Then, each group is placed at 4℃, 30℃, 50℃, 60℃, 70℃, 80℃, 90℃ and 100℃ respectively for 30 min. The enzyme activity of xylanase at different temperatures is calculated according to the enzyme activity assay method in Example 3.

[0061] Based on the above calculations, the enzyme activity at the measured optimal reaction temperature was used as 100% relative enzyme activity for plotting. The results are shown in Figure 3. It can be seen that the xylanase isolated in this invention has certain heat resistance, exhibiting high relative enzyme activity at high temperatures of 60~100℃, with an optimal reaction temperature of approximately 90℃.

[0062] (b) The optimal pH was referenced in the reaction systems of Example 3. The difference in this example is that the 0.1 M citrate buffer at pH 6.0 in each system was replaced with the corresponding buffers in the pH range of 3.5 to 10.5. Specifically, the buffers were: 0.1 M citrate buffers at pH 3.5, pH 4.5, pH 5.5, pH 6.0, and pH 6.5, and 0.1 M Tris-HCl buffers at pH 6.5, pH 7.5, pH 8.5, pH 9.5, and pH 10.5. Xylan was prepared with a mass concentration of 1% (w / v) using the buffers at the above different pH values ​​as the reaction substrate for the corresponding systems. The enzyme solution used was the purified xylanase solution diluted 5000 times. The reaction was carried out at 50°C for 30 min, and the enzyme activity of xylanase under different pH conditions was calculated according to the enzyme activity assay method in Example 3.

[0063] Based on the above calculations, the enzyme activity at the optimal pH was plotted as 100% relative enzyme activity. The results are shown in Figure 4. It can be seen that the xylanase isolated in this invention possesses certain alkali resistance, exhibiting high enzyme activity in the pH range of 6–10.5, with the optimal pH range being between 6 and 7.

[0064] (c) Thermal stability: Enzymes with thermal stability at high temperatures can maintain good enzyme activity in the extreme environments of industrial production, thus possessing higher industrial application value. Based on this, the thermal stability of the prepared xylanase was investigated in this embodiment.

[0065] Specifically, the xylanase purified solution diluted 5000 times was first subjected to incubation treatment for different durations (0 h, 0.5 h, 1 h, 1.5 h, 2 h, 2.5 h, 3 h) at different temperature gradients (50°C, 70°C, 100°C). Then, referring to the reaction systems in Example 3, the incubated enzyme was added to the enzyme reaction system and reacted in a 50°C water bath for 30 min. The enzyme activity of xylanase after different incubation treatments was calculated according to the enzyme activity assay method in Example 3.

[0066] Based on the above calculations, the highest enzyme activity under different temperature bath conditions was used as 100% relative enzyme activity for plotting. The results are shown in Figure 5. It can be seen that after bathing at 50℃ and 70℃ for 2 hours, the xylanase still retains approximately 80% of its activity, and after bathing at 100℃ for 1 hour, it still retains approximately 60% of its activity. This indicates that the xylanase possesses good thermostability and has good application potential in moderately extreme environments such as industrial production.

Claims

1. An application of a xylanase, characterized in that, The application includes: reacting xylanase with xylan or raw materials containing xylan under extreme conditions to achieve xylan hydrolysis; the extreme conditions include: temperature 60~100℃ and / or pH 6~10.5; the amino acid sequence of the xylanase is shown in SEQ ID NO.

1.

2. The application according to claim 1, characterized in that, The extreme environments include temperatures of 70–100°C and / or pH of 7–10.

5.

3. The application according to claim 1 or 2, characterized in that, The extreme environments include temperatures of 80–100°C and / or pH values ​​of 8–10.

5.

4. The application according to claim 3, characterized in that, The extreme environments include temperatures of 90-100°C and / or pH of 9-10.

5.

5. The application according to claim 4, characterized in that, The raw materials containing xylan include any one or more of the following: grain raw materials, agricultural waste, forestry raw materials, and textile raw materials.

6. A method for preparing xylooligosaccharides and / or xylose under extreme conditions, characterized in that, The method includes: adding xylanase with an amino acid sequence as shown in SEQ ID NO.1 to xylan or a raw material containing xylan, and carrying out a contact reaction at a temperature of 60~100℃ and / or pH 6~10.

5.

7. The method according to claim 6, characterized in that, The xylanase is added to xylan or raw materials containing xylan, and a contact reaction is carried out at a temperature of 90~100℃ and / or pH 9~10.

5.

8. The method according to claim 6 or 7, characterized in that, The raw materials containing xylan include any one or more of the following: grain raw materials, agricultural waste, forestry raw materials, and textile raw materials.

9. An enzyme preparation composition suitable for extreme environments, characterized in that, The composition includes a xylanase with the amino acid sequence shown in SEQ ID NO.1, which has enzymatic activity for catalyzing the hydrolysis of xylan at temperatures of 60-100°C and / or pH of 6-10.

5.

10. The enzyme preparation composition according to claim 9, characterized in that, The xylanase exhibits enzymatic activity that catalyzes the hydrolysis of xylan under conditions of temperature 90-100℃ and / or pH 9-10.5.

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