Structure of a serine-type glutathione S-transferase of Ceriporiopsis subvermispora and identification of the enzymatically important non-canonical residues by functional mutagenesis
Structure of a serine-type glutathione S-transferase of Ceriporiopsis subvermispora and identification of the enzymatically important non-canonical residues by functional mutagenesis
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DOI：
10.1016/j.bbrc.2019.01.076
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发表时间：
2019-02-26
影响因子：
3.1
通讯作者：
Katahira, Masato
中科院分区：
文献类型：
--
作者：
Osman, Wan Hasnidah Wan;Mikami, Bunzo;Katahira, Masato
枯草杆菌(Ceriporiopsis subvermispora，C.subvermispora)是一种白腐菌，是木质生物质的选择性木质素降解物。谷胱甘肽S转移酶(GSTs)是一种多功能的酶，能够催化参与解毒和代谢途径的反应。本研究在大肠杆菌中高效表达了枯草芽孢杆菌的GST，命名为CsGST63524，并通过亲和、阴离子交换和大小排斥柱层析对其进行了纯化。CsGST63524的晶体结构分别在2.45埃分辨率和2.50埃分辨率下进行了精细化。谷胱甘肽的硫原子与CsGST63524的Ser21形成氢键，表明它是丝氨酸型GST。Ser21的诱变意外地表明，该丝氨酸残基对CsGST63524的酶活性不是必需的。比较序列和结构分析，结合功能突变，新发现了除丝氨酸残基外的重要的非典型氨基酸残基Asn23和Tyr45。(C)2019 Elsevier Inc.保留所有权利。
Ceriporiopsis subvermispora (C. subvermispora), one of the white-rot fungi, is known as a selective lignin degrader of the woody biomass. Glutathione S-transferases (GSTs) are multifunctional enzymes that are capable of catalyzing the reactions involved in detoxification and metabolic pathways. In this study, a GST of C subvermispora, named CsGST63524, was overexpressed in E. coli, and then purified by affinity, anion exchange, and size exclusion column chromatography. The crystal structures of the CsGST63524 in ligand-free and complex with GSH were refined at 2.45 and 2.50 angstrom resolutions, respectively. The sulfur atom of glutathione forms a hydrogen bond with Ser21 of CsGST63524, indicating it is a serine-type GST. Mutagenesis of Ser21 unexpectedly indicated that this serine residue is not essential for the enzymatic activity of CsGST63524. Comparative sequence and structural analyses, together with functional mutagenesis, newly identified the enzymatically important non-canonical amino acid residues, Asn23 and Tyr45, other than the serine residue. (C) 2019 Elsevier Inc. All rights reserved.