Crystal Structure of F65A/Y131C-Methylimidazole Carbonic Anhydrase V Reveals Architectural Features of an Engineered Proton Shuttle

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• 作者：Kevin M. Jude ; S. Kirk Wright ; Chingkuang Tu ; David N. Silverman ; Ronald E. Viola ; and David W. Christianson
• 刊名：Biochemistry
• 出版年：2002
• 出版时间：February 26, 2002
• 年：2002
• 卷：41
• 期：8
• 页码：2485 - 2491
• 全文大小：260K
• 年卷期：v.41,no.8(February 26, 2002)
• ISSN：1520-4995

文摘

The crystal structure of F65A/Y131C murine -carbonic anhydrase V (CAV), covalentlymodified at cysteine residues with 4-chloromethylimidazole, is reported at 1.88 Å resolution. Thismodification introduces a methylimidazole (MI) group at residue C131 in the active site with importantconsequences. F65A/Y131C-MI CAV exhibits an up to 3-fold enhancement of catalytic activity over thatof wild-type CAV [Earnhardt, J. N., Wright, S. K., Qian, M., Tu, C., Laipis, P. J., Viola, R. E., andSilverman, D. N. (1999) Arch. Biochem. Biophys. 361, 264-270]. In this modified CAV variant, C131-MI acts as a proton shuttle, facilitating the deprotonation of a zinc-bound water molecule to regeneratethe nucleophilic zinc-bound hydroxide ion. A network of three hydrogen-bonded water molecules, acrosswhich proton transfer likely proceeds, bridges the zinc-bound water molecule and the C131-MI imidazolegroup. The structure of F65A/Y131C-MI CAV is compared to structures of Y64H/F65A murine CAV,wild-type human -carbonic anhydrase II, and the -carbonic anhydrase from Methanosarcina thermophilain an effort to outline common features of catalytic proton shuttles.