Isolation of a gene encoding endoglucanase activity from uncultured microorganisms in buffalo rumen

被引：18

作者：

Liu, Li ^{[1
,2
,3
]}

Feng, Yi ^{[1
,2
,4
]}

Duan, Cheng-Jie ^{[1
,2
]}

Pang, Hao ^{[1
,2
]}

Tang, Ji-Liang ^{[1
,2
]}

Feng, Jia-Xun ^{[1
,2
]}

机构：

[1] Guangxi Univ, Guangxi Key Lab Subtrop Bioresources Conservat &, Key Lab Minist Educ Microbial & Plant Genet Engn, Nanning 530005, Guangxi, Peoples R China

[2] Guangxi Univ, Coll Life Sci & Technol, Nanning 530005, Guangxi, Peoples R China

[3] Hezhou Univ, Hezhou 542800, Guangxi, Peoples R China

[4] Guangxi Univ Tradit Chinese Med, Nanning 530001, Guangxi, Peoples R China

来源：

WORLD JOURNAL OF MICROBIOLOGY & BIOTECHNOLOGY | 2009年 / 25卷 / 06期

基金：

中国国家自然科学基金;

关键词：

Endo-beta-1,4-glucanase; Expression; Characterization; umcel5N; Metagenomic library; Buffalo rumen; BETA-GLUCOSIDASE; CELLULASE; METAGENOME; GENOMICS; STRAIN;

D O I：

10.1007/s11274-009-9983-8

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

A gene, umcel5N, was isolated from a metagenomic library constructed from the contents of buffalo rumen. Its putative product belongs to the glycosyl hydrolase family 5 and is most closely related to an endoglucanase (ABN54006.1) from Clostridium thermocellum with 44% identity and 60% similarity. Gene umcel5N was heterologously expressed in Escherichia coli. The purified recombinant Umcel5N hydrolyzed carboxymethyl cellulose with a rapid decrease in the viscosity of the solution but with little release of reducing sugars, suggesting an endo mode of action. The enzyme exhibited optimal activity toward p-nitrophenyl beta-d-cellobioside at pH 5.5 and 55A degrees C, and had a Km of 1.56 mM and a Vmax of 285.6 U/mg. Two glutamic acids (E144 and E285) of the wild-type Umcel5N were predicted as a proton donor and a nucleophile, respectively. Site-directed mutagenesis confirmed that they were required for the enzyme's activity.

引用

页码：1035 / 1042

页数：8

共 15 条

[1] Cloning and identification of novel cellulase genes from uncultured microorganisms in rabbit cecum and characterization of the expressed cellulases [J].