CN102154246B - Acid glucanase CEL7G5 and gene and application thereof - Google Patents
Acid glucanase CEL7G5 and gene and application thereof Download PDFInfo
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- CN102154246B CN102154246B CN201110031673XA CN201110031673A CN102154246B CN 102154246 B CN102154246 B CN 102154246B CN 201110031673X A CN201110031673X A CN 201110031673XA CN 201110031673 A CN201110031673 A CN 201110031673A CN 102154246 B CN102154246 B CN 102154246B
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Abstract
The invention relates to the field of gene engineering and particularly relates to acid glucanase CEL7G5 and a gene and application thereof. The invention provides the glucanase CEL7G5 from Phialophora sp. An amino acid sequence of the acid glucanase CEL7G5 is shown as SEQ ID NO.1. The invention also provides a coding gene Cel7G5 of the glucanase. The glucanase has the following characteristics that the optimum pH (potential of hydrogen) is 5.0; the optimum temperature is 60 DEG C; the specific activity is 922.9U/mg; the glucanase has good proteinase resistance; by the glucanase, beta-glucan and hydroxymethyl cellulose are effectively degraded; and the glucanase is easy for industrial fermentation production. As a novel enzymic preparation, the acid glucanase CEL7G5 can be widely used for industries such as feeds, brewing, foods, energy and the like.
Description
Technical field
The present invention relates to the genetically engineered field, particularly, the present invention relates to a kind of acidic dextranase CEL7G5 and gene and application.
Background technology
Mierocrystalline cellulose, semicellulose and xylogen etc. are the main moitys of plant cell wall.Mierocrystalline cellulose accounts for 40～45% of dried cell weight greatly, is through β-1, the linear structure molecule that the 4-glycosidic link is formed by connecting by glucose.Xylogen accounts for 15～25% of dried cell weight, is a kind of complicated phenol polymer.Semicellulose accounts for 30～35% of dried cell weight, is renewable biological source the abundantest after Mierocrystalline cellulose.It is gathered polysaccharide and is formed by assorted, names by the sugar that the main chain composition occupies the majority, be referred to as VISOSE, VISOSE, mannosans and KGM etc. (Schulze E1891.Ber Dtsch Chem Ges 24,2277-2287.).Wherein beta-glucan is the structural non-starch polysaccharide in the unifacial leaf grass cell walls; Mainly be present in aleurone layer and the albuminous cell; As barley and oat albuminous cell wall contain 70～75% the VISOSE of having an appointment (Philippe S et al.2006.Planta 224 (2), 449-461.).
Beta-glucanase is can decompose β-glycosidic link chain to become the general name of the class of enzymes of glucose polymer.Different by the mode of action, beta-glucanase can be divided into two types of NCE5 and exoglucanases.Inscribe β-1 wherein, 3-1,4-LSD (zymetology classification number E.C.3.2.1.73) can hydrolysis β-1,3-1, the 4-VISOSE, single-minded β-1,4 glycosidic link that links to each other with β-1,3 key that acts on makes it be degraded to low molecular weight fraction, loses wetting ability and viscosity.Change the characteristic of monogastric animal intestinal contents; Improve the endogenous digestive enzyme activity; Change the enteric microorganism environment; Be beneficial to digestion and the absorption of animal to nutritive substance, (Mathlouthi N et al.2002.Amin Res's transformation efficiency of raising growth performance and feed 51 395-406.) has broad application prospects at aspects such as food and fodder industries.
Has the potential application prospect in the brewage industry; In the brewage process, the VISOSE in the Fructus Hordei Germinatus causes the beer filtration difficulty, stops up filtering membrane, has increased the production cost and the quality of beer; Adopt acidic dextranase and LSD synergy, can overcome the above problems.Therefore, the architecture basics of the generation of acidic dextranase, purifying, character, acidic character and the application in fields such as feed processing, wine industry, fruit juice processing and the energy thereof deepen continuously.
Because different industry are different to the LSD property requirements, therefore, obtain novel research with good characteristic LSD and still are significant.The clone with separate the LSD with high-temperature acidic can better application in feed, wine brewing, foodstuffs industry.LSD among the present invention all has the high enzyme vigor under acidity, neutral pH, have good pH stability and thermostability, and have higher cellulase activity, meets multiple industry needs.
Summary of the invention
The acidic dextranase LSD that the purpose of this invention is to provide a kind of ability efficient application.
A purpose more of the present invention provides the gene of the above-mentioned acidic dextranase of coding.
Another object of the present invention provides the recombinant vectors that comprises said gene.
Another object of the present invention provides the recombinant bacterial strain that comprises said gene.
Another object of the present invention provides a kind of gene engineering method for preparing above-mentioned acidic dextranase.
Another object of the present invention provides the application of above-mentioned acidic dextranase.
The present invention separates from Saksenaea vasiformis (Phialophora sp.) and obtains a kind of new acidic dextranase CEL7G5.
The invention provides a kind of acidic dextranase CEL7G5, its aminoacid sequence is shown in SEQ ID NO.1.SEQ?ID?NO.1：
MTAKFTALSALLGLAASQQVGTQQTETHPQMSWSKCSSGGSCTSQNGEVVIDANWRWVHDVGGYTNCYTGNEWNTTICPDDKTCAANCAVDGADYAATYGAPTSGNALTLKFVTKGSYLAPKSATNIGSRLYLMASSSKYQMFTLLGNEFTFDVDVSQLGCGLNGALYFVSMDADGGMSKYPTNKAGAKYGTGYCDSQCPRDLKFINGEANCDNWQPSSNDQNAGVGGYGSCCTEMDVWEANSISTAYTPHPCTTVGQERCTGDSCGGTYSSDRYGGECDPDGCDFNSYRMGVTNFYGPSMTVDTTQKFTVVTQFLTGSDGTLDEIKRFYVQGGQVIPNSNSTISGVTGNSITPDFCKAQKKAFGDGDYFDQKGGFPQFSKALAGEMVLVMSLWDDHYSNMLWLDSVYPTDASASEPGKARGTCATTSGVPADVESSQASDQVVYSNIKFGPIGSTFQEPSS
Wherein, 462 amino acid of this enzyme genes encoding, N holds 17 signal peptide sequences " mtakftalsallglaas " (SEQ ID NO.3) that amino acid is its prediction.
Therefore, the theoretical molecular of sophisticated acidic dextranase CEL7G5 is 47.5kDa, and its aminoacid sequence is shown in SEQ ID NO.2:
QQVGTQQTETHPQMSWSKCSSGGSCTSQNGEVVIDANWRWVHDVGGYTNCYTGNEWNTTICPDDKTCAANCAVDGADYAATYGAPTSGNALTLKFVTKGSYLAPKSATNIGSRLYLMASSSKYQMFTLLGNEFTFDVDVSQLGCGLNGALYFVSMDADGGMSKYPTNKAGAKYGTGYCDSQCPRDLKFINGEANCDNWQPSSNDQNAGVGGYGSCCTEMDVWEANSISTAYTPHPCTTVGQERCTGDSCGGTYSSDRYGGECDPDGCDFNSYRMGVTNFYGPSMTVDTTQKFTVVTQFLTGSDGTLDEIKRFYVQGGQVIPNSNSTISGVTGNSITPDFCKAQKKAFGDGDYFDQKGGFPQFSKALAGEMVLVMSLWDDHYSNMLWLDSVYPTDASASEPGKARGTCATTSGVPADVESSQASDQVVYSNIKFGPIGSTFQEPSS
LSD CEL7G5 of the present invention has good pH stability simultaneously, and in acid and neutral scope, all has characteristics such as high reactivity, protease inhibitor degraded under the normal temperature.LSD of the present invention, its optimum pH are 5.0, in the scope of pH2.0～9.0, keep the enzymic activity more than 60%; Optimum temperuture is 60 ℃; With stomach en-and trypsin treatment 60 minutes, enzymic activity maintained more than 85%.
The invention provides the above-mentioned acidic dextranase Cel7G5 of coding.Particularly, the genome sequence of this gene is shown in SEQ ID NO.4:
atgactgccaaattcactgccctttcagcccttctgggccttgccgcttcgcagcaagtgggcacccagcaaaccgagacccatccgcagatgtcctggtccaaatgctctagcggaggctcctgcaccagccagaacggcgaggttgtgatcgacgccaactggcgttgggtccatgacgttggaggctacaccaactgctacaccggcaacgagtggaacaccaccatctgccccgacgacaagacctgtgctgccaactgtgctgtggacggcgccgactacgcggctacatatggtgccccgaccagcggcaacgccctgaccctcaagttcgtgaccaagggctcgtatttagcgcccaaatcagcgaccaacattgggtcgcgtctctacctgatggccagctccagcaagtaccagatgttcaccttgctgggcaacgagttcaccttcgatgtcgatgtctcccagctcggctgcggtctcaacggcgctctttacttcgtgtccatggacgcggacggcggcatgtccaagtatcccaccaacaaggccggagccaagtacggaaccggctactgcgactctcagtgcccgcgtgatctgaagttcatcaacggtgaggccaactgtgataactggcagccatcctcgaacgaccagaacgccggtgttggtggatatggttcctgctgtactgagatggacgtctgggaggccaactcgatctcgacggcctacactccccacccctgcaccacggtgggccaggagcgctgcacgggcgattcctgtggaggaacgtattcttcggaccgctacggcggtgaatgcgatccggacggctgtgacttcaacagctaccgcatgggtgtcaccaacttctacggccccagcatgacggtcgataccacccagaagttcaccgtcgtgactcagttcctcacgggcagcgacggcacactggacgagatcaagcgcttctatgtccagggtggccaggtgatccccaactccaactccaccatctccggcgtcacgggcaactccatcacccccgacttctgcaaggctcagaagaaggccttcggcgacggagactacttcgaccagaagggaggcttcccccagttcagcaaggctctcgccggggagatggtgctcgtcatgtccctctgggatgaccactactccaacatgctctggctcgactccgtgtacccgaccgacgcctctgccagtgaacccggcaaggcccgcggtacctgcgctaccacatcgggtgtccctgccgacgtcgagtccagccaagccagcgaccaggtcgtctactcgaacatcaagttcggccccatcggctcgaccttccaggaaccgtcgtcctaa
The method separating clone of the present invention through PCR glucanase gene Cel7G5, the DNA complete sequence analysis is the result show, LSD CEL7G5 structure gene Cel7G5 total length 1389bp.Wherein, the base sequence of signal peptide is:
ATGACTGCCAAATTCACTGCCCTTTCAGCCCTTCTGGGCCTTGCCGCTTCG(SEQ?ID?NO.6)。
The gene order of sophisticated LSD CEL7G5 is shown in SEQ ID NO.5.
SEQ?ID?NO.5
cagcaagtgggcacccagcaaaccgagacccatccgcagatgtcctggtccaaatgctctagcggaggctcctgcaccagccagaacggcgaggttgtgatcgacgccaactggcgttgggtccatgacgttggaggctacaccaactgctacaccggcaacgagtggaacaccaccatctgccccgacgacaagacctgtgctgccaactgtgctgtggacggcgccgactacgcggctacatatggtgccccgaccagcggcaacgccctgaccctcaagttcgtgaccaagggctcgtatttagcgcccaaatcagcgaccaacattgggtcgcgtctctacctgatggccagctccagcaagtaccagatgttcaccttgctgggcaacgagttcaccttcgatgtcgatgtctcccagctcggctgcggtctcaacggcgctctttacttcgtgtccatggacgcggacggcggcatgtccaagtatcccaccaacaaggccggagccaagtacggaaccggctactgcgactctcagtgcccgcgtgatctgaagttcatcaacggtgaggccaactgtgataactggcagccatcctcgaacgaccagaacgccggtgttggtggatatggttcctgctgtactgagatggacgtctgggaggccaactcgatctcgacggcctacactccccacccctgcaccacggtgggccaggagcgctgcacgggcgattcctgtggaggaacgtattcttcggaccgctacggcggtgaatgcgatccggacggctgtgacttcaacagctaccgcatgggtgtcaccaacttctacggccccagcatgacggtcgataccacccagaagttcaccgtcgtgactcagttcctcacgggcagcgacggcacactggacgagatcaagcgcttctatgtccagggtggccaggtgatccccaactccaactccaccatctccggcgtcacgggcaactccatcacccccgacttctgcaaggctcagaagaaggccttcggcgacggagactacttcgaccagaagggaggcttcccccagttcagcaaggctctcgccggggagatggtgctcgtcatgtccctctgggatgaccactactccaacatgctctggctcgactccgtgtacccgaccgacgcctctgccagtgaacccggcaaggcccgcggtacctgcgctaccacatcgggtgtccctgccgacgtcgagtccagccaagccagcgaccaggtcgtctactcgaacatcaagttcggccccatcggctcgaccttccaggaaccgtcgtcctaa
The maturation protein theoretical molecular is 47.5kDa; Glucanase gene Cel7G5 sequence and the aminoacid sequence derived are carried out the BLAST comparison in GenBank, this gene is 71% with the cellulase consensus amino acid sequence property that derives from Acremonium thermophilum.Explain that CEL7G5 is a kind of new LSD.
The present invention also provides the recombinant vectors that comprises above-mentioned acidic dextranase gene C el7G5, is preferably pPIC-Cel7G5.Glucanase gene of the present invention is inserted between the suitable restriction enzyme site of expression vector, makes that its nucleotide sequence is exercisable to be connected with expression regulation sequence.As a most preferred embodiment of the present invention; Be preferably glucanase gene of the present invention is inserted between the EcoR I and Not I restriction enzyme site on the plasmid pPIC9; Make this nucleotide sequence be positioned at the downstream of AOX1 promotor and regulated and control by it, obtain expression of recombinant yeast plasmid pPIC9-Cel7G5.
The present invention also provides the recombinant bacterial strain that comprises above-mentioned acidic dextranase gene C el7G5, and preferred said bacterial strain is intestinal bacteria, yeast, genus bacillus or lactobacillus spp, is preferably recombinant bacterial strain GS115/Cel7G5.
The present invention also provides a kind of method for preparing acidic dextranase CEL7G5, may further comprise the steps:
1), gets recombinant bacterial strain with above-mentioned recombinant vectors transformed host cell;
2) cultivate recombinant bacterial strain, induce the reorganization LSD to express; And
3) reclaim the also expressed LSD CEL7G5 of purifying.
Wherein, preferred said host cell is pichia spp cell, cerevisiae or many types of inferior yeast cell, preferably the expression of recombinant yeast plasmid is transformed pichia spp cell (Pichia pastoris) GS115, obtains recombinant bacterial strain GS115/Cel7G5.
The present invention also provides the application of above-mentioned acidic dextranase CEL7G5.
The present invention's technical problem at first to be solved is the deficiency that overcomes prior art, provide a kind of character good, be suitable at feed, wine brewing, LSD that Applications in Food Industry is new.LSD ph optimum of the present invention is 5.0, in pH2.0～6.0 higher enzymic activity is arranged all; The pH good stability; Ability with good protease inhibitor; And has higher dextranase activity.This LSD can be applicable to fodder industry, effectively reduces viscosity, and elimination or reduction increase the anti-oxidant action that causes because of viscosity.In wine industry, the VISOSE of can effectively degrading, the viscosity that effectively reduces wort improves the filtration efficiency clarifying beer.Therefore, the application of this LSD in energy industry also demonstrates its great potential.
Description of drawings
The recombinate ph optimum of LSD of Fig. 1.
The recombinate pH stability of LSD of Fig. 2.
The recombinate optimum temperuture of LSD of Fig. 3.
The recombinate thermostability of LSD of Fig. 4.
Embodiment
Test materials and reagent
1, bacterial strain and carrier: the present invention separates from Saksenaea vasiformis (Phialophora sp.) and obtains a kind of new acidic dextranase CEL7G5.Yeast expression vector pPIC9 and bacterial strain GS115 are available from Invitrogen company.
2, enzyme and other biochemical reagents: restriction endonuclease is available from TaKaRa company, and ligase enzyme is available from Invitrogen company.Oat beta-glucan is available from Sigma company, and other all is domestic reagent (all can buy from common biochemical reagents company and obtain).
3, substratum:
(1) Phialophora sp. substratum is the potato juice substratum: 1000mL potato juice, 10g glucose, 25g agar, pH2.5.
(2) intestinal bacteria substratum LB (1% peptone, 0.5% yeast extract, 1%NaCl, pH7.0).
(3) BMGY substratum: 1% yeast extract, 2% peptone, 1.34%YNB, 0.00004%Biotin, 1% glycerine (V/V).
(4) BMMY substratum: replace glycerine divided by 0.5% methyl alcohol, all the other compositions are all identical with BMGY, pH4.0.
Explain: make the experimental methods of molecular biology specify in following examples, all carry out, perhaps carry out according to test kit and product description with reference to listed concrete grammar in " molecular cloning experiment guide " (third edition) J. Sa nurse Brooker one book.
The clone of embodiment 1 Saksenaea vasiformis Phialophora sp. glucanase coding gene as well as Cel7G5
Extract Saksenaea vasiformis Phialophora sp. genomic dna:
3 days mycelium of liquid culture is put into mortar with the aseptic filter paper filtration, add the 2mL extracting solution, grind 5min; Then lapping liquid is placed the 50mL centrifuge tube; 65 ℃ of water-bath cracking 20min, whenever once at a distance from the 10min mixing, at 4 ℃ of centrifugal 5min of following 10000rpm.Get supernatant extrct foreigh protein removing in phenol/chloroform, get supernatant again and add the equal-volume Virahol, after room temperature leaves standstill 5min, 4 ℃ of centrifugal 10min of following 10000rpm.Abandon supernatant, deposition is with 70% washing with alcohol twice, and vacuum-drying adds an amount of TE and dissolves, place-20 ℃ subsequent use.
Conservative (GYCDA (S) QC and GCG (D) FNPY) sequences Design according to the 7th family's glucanase gene has been synthesized degenerated primer P1, P2
P1：5′-GGYTACTGYGAYGCNCARTG-3′；
P2：5′-TAGGGRTTGAANCCRCANCC-3′)。
With the total DNA of Phialophora sp. is that template is carried out pcr amplification.The PCR reaction parameter is: 94 ℃ of sex change 5min; 94 ℃ of sex change 30sec then, 48 ℃ of annealing 30sec, 72 ℃ are extended 1min, 30 back 72 ℃ of insulation 10min of circulation.Obtain an about 291bp fragment, this fragment recovery back is linked to each other with the pEASY-T3 carrier send the order-checking of three rich Bioisystech Co., Ltd.
According to the nucleotide sequence that order-checking obtains, each three TAIL-PCR Auele Specific Primer of design upstream and downstream: design direction is for needing the zone of ignorance direction of amplification, and the Position Design of sp2 is in the inboard of sp1, and sp3 is positioned at the inboard of sp2.Distance between per two primers does not have strict regulation, the general 22～30nt of primer length, and annealing temperature is at 60～65 ℃.
Table 1. LSD CEL7G5TAIL-PCR Auele Specific Primer
Obtain the flanking sequence of known sequence through TAIL-PCR, amplification obtains sending after product reclaims the order-checking of three rich Bioisystech Co., Ltd.Splicing back CEL7G5 glucanase gene total length 1389bp, encode 462 amino acid and a terminator codon.Carry out the signal peptide of 17 amino acid of analysis revealed N end with SignalP (http://www.cbs.dtu.dk/services/SignalP) for prediction.The theoretical molecular of predicting the maturation protein of this coded by said gene is 47.5kDa.
The preparation of embodiment 2 reorganization LSDs
Expression vector pPIC9 is carried out double digestion (EcoR I+Not I); To encode the simultaneously gene C el7G5 double digestion (EcoR I+Not I) of LSD; The gene fragment that cuts out the encoding mature LSD is connected with expression vector pPIC9; Acquisition contains the recombinant plasmid pPIC-Cel7G5 of Phialophora sp. glucanase gene Cel7G5 and transforms pichia spp GS115, obtains recombinant pichia yeast strain GS115/Cel7G5.
Get the GS115 bacterial strain that contains recombinant plasmid, be inoculated in the 300mL BMGY nutrient solution, behind 30 ℃ of 250rpm shaking culture 48h, centrifugal collection thalline.Resuspended in 150mL BMMY substratum then, 30 ℃ of 250rpm shaking culture.After inducing 72h, centrifugal collection supernatant.Measure the vigor of LSD.The expression amount of reorganization LSD is 21.6U/mL.SDS-PAGE result shows that the reorganization LSD has obtained expression in pichia spp.The ratio of reorganization VISOSE is lived and is 922.9U/mg.
The activation analysis of embodiment 3 reorganization LSDs
The DNS method: concrete grammar is following: at pH5.0, under 60 ℃ of conditions, the reaction system of 1mL comprises 100 μ L suitable dilution enzyme liquid, 900 μ L substrates, and reaction 10min adds the 1.5mLDNS termination reaction, and boiling water boils 5min.Cooling back 540nm measures the OD value.1 enzyme unit (U) that lives is defined as the enzyme amount that under given condition PM discharges 1 μ mol reducing sugar.
The property testing of embodiment 4 reorganization LSD CEL7G5
1, the measuring method of the ph optimum of reorganization LSD CEL7G5 and pH stability is following:
2, the optimum temperuture of LSD and thermal stability determination method are following:
Enzymatic reaction is carried out in being determined as under Hydrocerol A-Sodium phosphate, dibasic damping fluid (pH5.0) buffer solution system and differing temps of the optimum temperuture of LSD.Temperature tolerance is determined as LSD and under differing temps, handles different time, under 60 ℃, carries out enzyme assay again.The enzyme reaction optimum temperuture is measured result (Fig. 3) and is shown that its optimum temperuture is 60 ℃.The thermostability property test of enzyme shows (Fig. 4), and CEL7G5 has good thermostability, at 55 ℃ of following incubation 2h, can keep the enzyme more than 90% to live.
3, the K of LSD
mValues determination method is following:
Use the VISOSE of different concns to be substrate, in Hydrocerol A-Sodium phosphate, dibasic damping fluid (pH4.0) buffer solution system, measure enzymic activity down, calculate its K for 60 ℃
mValue.Through measuring the K when being substrate with VISOSE and sodium cellulose glycolate
mValue is respectively 2.7 and 3268.0mg/mL, maximum reaction velocity V
MaxBe respectively 2.2 and 826.6 μ mol/minmg.
4, different metal ion chemistry reagent is measured as follows the influence of CEL7G5 enzyme work:
In enzymatic reaction system, add the different metallic ion and the chemical reagent of different concns, study its influence to enzymic activity, various material final concentrations are 1 and 5mmol/L.Under 60 ℃, pH5.0 condition, measure enzymic activity.The result shows, most of ions and chemical reagent do not have considerable change at the vigor of concentration reorganization LSD during for 1mmol.But Ag
+, Hg
2+Almost can suppress its vigor fully, and also its vigor of strongly inhibited of SDS.Work as Cu
2+, pb
2+, EDTA partly suppresses its enzyme activity when concentration is 10mmol.And beta-mercaptoethanol can partly activate the CEL7G5 enzyme activity.
5, LSD antipepsin and trypsinase ability are measured as follows:
With pH2.0KCl-HCl damping fluid preparation 0.1mg/mL stomach en-, pH7.0Tris-HCl damping fluid preparation 0.1mg/mL trypsinase.The enzyme liquid of getting the 0.5mL purifying after the pH2.0KCl-HCl damping fluid dilutes adds the 0.5mL stomach en-; The enzyme liquid of the 0.5mL purifying after the dilution of pH7.0Tris-HCl damping fluid adds 0.5mL trypsinase and mixes; Proteolytic enzyme/LSD (w/w) ≈ 0.1; 37 ℃ of insulations 30 are taken a sample with 60min, under pH5.0 and 60 ℃ of conditions, measure enzymic activity.After experimental result showed that LSD CEL7G5 is with stomach en-and trypsin treatment 60min, the enzyme that the CEL7G5 after the protease treatment still has more than 85% was lived.
6, the substrate specificity of reorganization LSD
This enzyme also has certain Degradation except that acting on the VISOSE for sodium cellulose glycolate.It is 29% to the degradation capability of sodium cellulose glycolate with respect to VISOSE.
7, add the influence of zytase to barley germ juice viscosity and filtration velocity
Barley germ is handled through kibbler, crosses the 0.2mm screen cloth, is dissolved in 100mL Hydrocerol A-Sodium phosphate, dibasic damping fluid.The reorganization LSD Cel7G5 of interpolation 40 or 80U.Handle 30min at 45,50 ℃ respectively then, handle 60min for 60 ℃, handle 30min, boil the 5min deactivation at last for 70 ℃.Experiment contrast is not for adding LSD.Measure its filtration velocity with filter paper.Get the 5ml filtered liq and measure its viscosity numerical value with the viscosity agent.The result shows, add the enzyme liquid of 50U and handle, its with compare, filtration velocity and viscosity reduce by 18.4% and 6.1% respectively.
Claims (10)
1. an acidic dextranase CEL7G5 is characterized in that, its aminoacid sequence is shown in SEQ ID NO.1 or SEQ ID NO.2.
2. acidic dextranase CEL7G5 as claimed in claim 1 is characterized in that, sequence SEQ ID NO.1 comprises signal peptide at the N end, and the sequence of said signal peptide is shown in SEQ ID NO.3.
3. an acidic dextranase gene C el7G5 is characterized in that, the described acidic dextranase CEL7G5 of coding claim 1.
4. acidic dextranase gene C el7G5 as claimed in claim 3 is characterized in that, its base sequence is shown in SEQ ID NO.4 or SEQ ID NO.5.
5. acidic dextranase gene C el7G5 as claimed in claim 3 is characterized in that, said acidic dextranase gene C el7G5 comprises signal peptide sequence, and said sequence is shown in SEQ ID NO.6.
6. the recombinant vectors that comprises the said acidic dextranase gene C of claim 3 el7G5.
7. the recombinant vectors pPIC-Cel7G5 that comprises the said acidic dextranase gene C of claim 3 el7G5.
8. the recombinant bacterial strain that comprises the said acidic dextranase gene C of claim 3 el7G5.
9. a method for preparing acidic dextranase gene C el7G5 is characterized in that, may further comprise the steps:
1) with the recombinant vectors transformed host cell of claim 6, gets recombinant bacterial strain;
2) cultivate recombinant bacterial strain, induce the reorganization LSD to express;
3) reclaim the also expressed LSD CEL7G5 of purifying.
10. the said acidic dextranase CEL7G5 of claim 1 application of VISOSE that is used to degrade.
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