Structural Insights into the Dehydroascorbate Reductase Activity of Human Omega-Class Glutathione Transferases
- 作者：Huina Zhou<sup>1sup><sup> ; sup><sup>joan_zhn@yahoo.com.cnsup> ; Joseph Brock<sup>2sup><sup> ; sup><sup>joeylives2ride@gmail.comsup> ; Dan Liu<sup>1sup><sup> ; sup><sup>dan.liu@anu.edu.ausup> ; Philip G. Board<sup>1sup><sup> ; sup><sup>Philip.Board@anu.edu.ausup> ; Aaron J. Oakley<sup>2sup><sup> ; sup><sup>3sup><sup> ; sup><sup>aarono@uow.edu.ausup>
- 关键词：GST ; glutathione transferase ; GSH ; glutathione ; AA ; ascorbic acid ; DHA ; dehydroascorbate ; DHAR ; DHA reductase ; PDB ; Protein Data Bank ; MPD ; 2-methyl-2 ; 4-pentanediol ; PEG ; polyethylene glycol
- 刊名：Journal of Molecular Biology
- 出版年：2012
- 期刊代码：102_00222836
- 类别：imm
- 出版时间：13 July, 2012
- 卷：420
- 期：3
- 页码：190-203
- 文件大小：2090 K
摘要
The reduction of dehydroascorbate (DHA) to ascorbic acid (AA) is a vital cellular function. The omega-class glutathione transferases (GSTs) catalyze several reductive reactions in cellular biochemistry, including DHA reduction. In humans, two isozymes (GSTO1-1 and GSTO2-2) with significant DHA reductase (DHAR) activity are found, sharing 64%sequence identity. While the activity of GSTO2-2 is higher, it is significantly more unstable in vitro. We report the first crystal structures of human GSTO2-2, stabilized through site-directed mutagenesis and determined at 1.9聽脜 resolution in the presence and absence of glutathione (GSH). The structure of a human GSTO1-1 has been determined at 1.7聽脜 resolution in complex with the reaction product AA, which unexpectedly binds in the G-site, where the glutamyl moiety of GSH binds. The structure suggests a similar mode of ascorbate binding in GSTO2-2. This is the first time that a non-GSH-based reaction product has been observed in the G-site of any GST. AA stacks against a conserved aromatic residue, F34 (equivalent to Y34 in GSTO2-2). Mutation of Y34 to alanine in GSTO2-2 eliminates DHAR activity. From these structures and other biochemical data, we propose a mechanism of substrate binding and catalysis of DHAR activity.