Catalytic efficiency of HAP phytases is determined by a key residue in close proximity to the active site

文献类型： 外文期刊

第一作者： Fu, Dawei

作者： Fu, Dawei;Li, Zhongyuan;Huang, Huoqing;Yuan, Tiezheng;Shi, Pengjun;Luo, Huiying;Meng, Kun;Yang, Peilong;Yao, Bin

作者机构：

关键词： Escherichia coli;Phytase;Site-directed mutagenesis;Specific activity;Yersinia enterocolitica

期刊名称：APPLIED MICROBIOLOGY AND BIOTECHNOLOGY （ 影响因子：4.813； 五年影响因子：4.697 ）

ISSN：

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收录情况： SCI

摘要： The maximum activity of Yersinia enterocolitica phytase (YeAPPA) occurs at pH 5.0 and 45 °C, and notably, its specific activity (3.28±0.24 Umg-1) is 800-fold less than that of its Yersinia kristeensenii homolog (YkAPPA; 88% amino acid sequence identity). Sequence alignment and molecular modeling show that the arginine at position 79 (Arg79) in YeAPPA corresponding to Gly in YkAPPA as well as other histidine acid phosphatase (HAP) phytases is the only non-conserved residue near the catalytic site. To characterize the effects of the corresponding residue on the specific activities of HAP phytases, Escherichia coli EcAPPA, a well-characterized phytase with a known crystal structure, was selected for mutagenesis-its Gly73 was replaced with Arg, Asp, Glu, Ser, Thr, Leu, or Tyr. The results show that the specific activities of all of the corresponding EcAPPA mutants (17-2,400 Umg -1) were less than that of the wild-type phytase (3,524 Umg -1), and the activity levels were approximately proportional to the molecular volumes of the substituted residues' side chains. Site-directed replacement of Arg79 in YeAPPA (corresponding to Gly73 of EcAPPA) with Ser, Leu, and Gly largely increased the specific activity, which further verified the key role of the residue at position 79 for determining phytase activity. Thus, a new determinant that influences the catalytic efficiency of HAP phytases has been identified.

分类号： Q939.9

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