[1]贺淹才,刘爱花,张荣奎,等.嗜麦芽窄食单胞菌产生的几丁质酶的特性[J].华侨大学学报(自然科学版),2008,29(2):245-249.[doi:10.11830/ISSN.1000-5013.2008.02.0245]
　HE Yan-cai,LIU Ai-hua,ZHANG Rong-kui,et al.Characterization of a Chitinase from Stenotrophomonas maltophilia[J].Journal of Huaqiao University(Natural Science),2008,29(2):245-249.[doi:10.11830/ISSN.1000-5013.2008.02.0245]
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嗜麦芽窄食单胞菌产生的几丁质酶的特性()
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《华侨大学学报（自然科学版）》[ISSN:1000-5013/CN:35-1079/N]

卷:
第29卷
期数:
2008年第2期
页码:
245-249
栏目:
出版日期:
2008-04-20

文章信息/Info

Title:
Characterization of a Chitinase from Stenotrophomonas maltophilia
文章编号:
1000-5013(2008)02-0245-05
作者:
贺淹才; 刘爱花; 张荣奎; 刘治江
华侨大学材料科学与工程学院; 华侨大学材料科学与工程学院 福建泉州362021; 福建泉州362021
Author(s):
HE Yan-cai;  LIU Ai-hua;  ZHANG Rong-kui;  LIU Zhi-jiang
College of Material Science and Engineering, Huaqiao University, Quanzhou 362021, China
关键词:
几丁质酶;  嗜麦芽窄食单胞菌;  发酵培养;  分子结构;  生物信息学
Keywords:
chitinase;  Stenotrophomonas maltophilia;  fermentation culture;  molecular structure;  bioinformatics
分类号:
Q936
DOI:
10.11830/ISSN.1000-5013.2008.02.0245
文献标志码:
A
摘要:
从土壤中筛选出一株产几丁质酶的革兰氏阴性细菌,通过16S rDNA法对酶活力最高的菌株C进行鉴定,确定为嗜麦芽窄食单胞菌(S.maltophilia)菌株.通过单因素优化法和均匀设计法确定S.maltophilia产几丁质酶的最佳条件:在pH值为7.0的基础培养基中添加质量分数为1.0%的酵母膏、质量分数为0.5%的胶体几丁质,于180 r.min-1,30℃的摇床中培养60 h.对硫酸铵沉淀获得的粗酶进行酶学性质研究,结果表明,该几丁质酶的最适反应温度为50℃,最适反应pH值为7.2,在55℃以上的温度条件下容易失活.通过聚丙烯酰胺凝胶电泳分析,确定S.maltophilia几丁质酶的相对分子质量为6.8×103; 以筛选的S.malto-philia菌株的总DNA为模板,利用聚合酶链式反应(PCR)扩增出几丁质酶DNA测序; 应用生物信息学手段推导S.maltophilia几丁质酶为分泌型蛋白酶,预测其等电点pI值为5.42,相对分子质量为6.8×103(与实验纯化的酶蛋白电泳结果一致).预测该酶氨基酸序列具有信号肽区(氨基酸残基1~41)、Ⅲ型几丁质结合区(残基47~92)、多囊肾病域(107~194)、类纤维连接蛋白Ⅲ型区(Fn3,201~278)和18家族糖基水解域等5个结构功能区,5个区域被富含Ala,Gly,Pro,Ser和Thr的短序列连接起来; 在第400~500个氨基酸残基间有一个螺旋结构.
Abstract:
A strain of Stenotrophomonas maltophilia which produced extacellular chitinase was isolated from chitin-rich soils of shrimp drying fields.The fermentation experiments showed that the suitable chitinase producing media were containing 1.5% colloid chitin and 1.0% yeast extract,and cultured for 60 hours(180 r·min-1) in shake flask.The optimal pH and temperature were 7.0 and 30 ℃,respectively.The crude enzyme was obtained from the fermentation broth by ammonium sulfate precipitation process,the enzyme optimal temperature was 50 ℃ and pH was 7.2,respectively.The stability of chitinase decreased rapidly when the temperature get above 55 ℃.The chitinase was 6.8×103 by SDS-PAGE analysis.Some properties of this enzyme were deduced by bioinformatics method: this is a secretary protease; its molecular mass is 6.8×103(consistent with the result by extraction chitinase electrophoresis).The predicted pI is 5.42.The enzyme amino acid sequence has five domains: signal peptide(residues 1~41),type Ⅲ chitin-binding domain(ChtBD3)(residues 47~92),polycystic kidney disease(PKD) domain(residues 107~194),fibronectin type Ⅲ(Fn3) domain(residues 201~278),and family 18 glycosyl hydrolases(Glyco-18) catalytic domain.Spanning each of the five domains are short sequences full of Gly,Ala,Pro,Ser,and Thr.There is a helix structure between 400th~500th residues.

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备注/Memo

备注/Memo:
福建省自然科学基金资助项目(C04010011)
更新日期/Last Update: 2014-03-23