Gene cloning, expression, and characterization of a novel beta-mannanase from Bacillus circulans CGMCC 1416
文献类型: 外文期刊
第一作者: Li, Yanan
作者: Li, Yanan;Yang, Peilong;Meng, Kun;Wang, Yarn;Luo, Huiying;Yao, Bin;Li, Yanan;Wu, Ningfeng;Fan, Yunliu
作者机构:
关键词: beta-mannanase;Bacillus circulans;gene expression;characterization
期刊名称:JOURNAL OF MICROBIOLOGY AND BIOTECHNOLOGY ( 影响因子:2.351; 五年影响因子:2.65 )
ISSN: 1017-7825
年卷期: 2008 年 18 卷 1 期
页码:
收录情况: SCI
摘要: A DNA fragment containing 2,079 base pairs from Bacillus circulans CGMCC 1416 was cloned using degenerate PCR and inverse PCR. An open reading frame containing 981 bp was identified that encoding 326 amino acids residues, including a putative signal peptide of 31 residues. The deduced amino acid sequence showed the highest identity (68.1 %) with endo-beta-1,4-D-mannanase from Bacillus circulans strain K-1 of the glycoside hydrolase family 5 (GH5). The sequence encoding the mature protein was cloned into the pET-22b(+) vector and expressed in Escherichia coli as a recombinant fusion protein containing an N-terminal hexahistidine sequence. The fusion protein was purified by Ni2+ affinity chromatography and its hexahistidine tag cleaved to yield a 31-kDa beta-mannanase having a specific activity of 481.55 U/mg. The optimal activity of the purified protein, MANB48, was at 58 degrees C and pH 7.6. The hydrolysis product on substrate locust bean gum included a monosaccharide and mainly oligosaccharides. The recombinant MANB48 may be of potential use in the feed industry.
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