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A Novel β-mannanase with High Specific Activity from Bacillus circulans CGMCC1554: Gene Cloning, Expression and Enzymatic Characterization

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Abstract

A DNA fragment of 2,042 bp containing a novel β-mannanase gene, man5A, was identified from the genome of the mannan-degrading bacterium Bacillus circulans CGMCC1554. The open reading frame of man5A comprised 978 bp encoding a protein of 326 amino acids with a predicted molecular weight of 32 kDa. The amino acid sequence of the encoded mannanase, MAN5A, showed the highest identity (78.5%) to β-mannanases belonging to glycosyl hydrolases family 5. The gene man5A was efficiently expressed in Escherichia coli and Pichia pastoris with the highest activity of 541 U/ml in a 3-L fermenter. Recombinant MAN5A purified from E. coli had a high specific activity of 4,839 U/mg, which is much higher than that of enzymes that showed high sequence identity. The enzyme showed maximum activity at pH 7.6 and 60 °C and resistance to trypsin. After hydrolysis of LBG, oligomannosides accounted for 76% of the hydrolysis products. All these properties collectively make MAN5A a better candidate than current mannanases for use in the food and feed industry.

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Acknowledgments

This work was supported by the Chinese National High Technology Research and Development Program (863 Program, Grant No. 2007AA100601) and the Chinese Agricultural Microorganism Collection and Share Program (No. 2005DKA21201).

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Authors and Affiliations

  1. Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, 100081, China

    Peilong Yang, Yanan Li, Yaru Wang, Kun Meng, Huiying Luo, Tiezheng Yuan, Yingguo Bai & Bin Yao

  2. Department of Biochemistry, Guangzhou Medical College, Guangzhou, 510182, China

    Yanan Li

  3. SUNHY Group, Wuhan, 430007, China

    Zhichun Zhan

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  1. Peilong Yang
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  2. Yanan Li
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Correspondence to Bin Yao.

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Yang, P., Li, Y., Wang, Y. et al. A Novel β-mannanase with High Specific Activity from Bacillus circulans CGMCC1554: Gene Cloning, Expression and Enzymatic Characterization. Appl Biochem Biotechnol 159, 85–94 (2009). https://doi.org/10.1007/s12010-008-8364-3

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  • DOI: https://doi.org/10.1007/s12010-008-8364-3

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