Preparation, purification, crystallization and preliminary crystallographic analysis of dual-domain beta-propeller phytase from Bacillus sp HJB17

文献类型： 外文期刊

第一作者： Lu, Fang

作者： Lu, Fang;Guo, Gangxin;Li, Qianqian;Feng, Duo;Liu, Yong;Gao, Wei;Huang, Huoqing;Yang, Peilong;Gao, Wei

作者机构：

期刊名称：ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY COMMUNICATIONS （ 影响因子：1.056； 五年影响因子：1.089 ）

ISSN： 2053-230X

年卷期： 2014 年 70 卷

页码：

收录情况： SCI

摘要： beta-Propeller phytases (BPPs) are abundant in nature. Recently, dual-domain BPPs have been found in which the typical BPP domain is responsible for phytate hydrolysis. The dual-domain BPP (PhyH) from Bacillus sp. HJB17 was obtained with an incomplete N-terminal BPP domain (PhyH-DI; residues 41-318) and a typical BPP domain (PhyH-DII; residues 319-644) at the C-terminus. PhyH-DI was found to act synergistically (with a 1.2-2.5-fold increase in phosphate release) with PhyH-DII, other BPPs (PhyP and 168PhyA) and a histidine acid phosphatase. The structure of PhyH was therefore studied with the aim of explaining these functions. PhyH with the secreted signal peptide of the first 40 amino acids deleted (PhyHT) was cloned and expressed in Escherichia coli. Purified and active PhyHT protein was obtained by refolding from the precipitant. PhyHTwas crystallized using the vapour-diffusion method. The crystal grew in a condition consisting of 0.2 M sodium acetate trihydrate, 0.1 M Tris-HCl pH 9.5, 25%(w/v) polyethylene glycol 4000 using 1 mg ml(-1) protein solution at 289 K. A complete data set was collected from a crystal to 2.85 angstrom resolution using synchrotron radiation at 100 K. The crystal belonged to space group P12(1)1, with unit-cell parameters a = 46.82, b = 140.19, c = 81.94 angstrom, alpha = 90.00, beta = 92.00, gamma = 90.00 degrees. The asymmetric unit was estimated to contain one molecule of PhyHT.

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