Evolved Stereoselective Hydrolases for Broad-Spectrum G-Type Nerve Agent Detoxification

被引：82

作者：

Goldsmith, Moshe ^{[1
]}

Ashani, Yacov ^{[2
,4
]}

Simo, Yair ^{[1
]}

Ben-David, Moshe ^{[2
]}

Leader, Haim ^{[3
]}

Silman, Israel ^{[4
]}

Sussman, Joel L. ^{[2
]}

Tawfik, Dan S. ^{[1
]}

机构：

[1] Weizmann Inst Sci, Dept Biol Chem, IL-76100 Rehovot, Israel

[2] Weizmann Inst Sci, Dept Biol Struct, IL-76100 Rehovot, Israel

[3] Weizmann Inst Sci, Dept Mat & Interfaces, IL-76100 Rehovot, Israel

[4] Weizmann Inst Sci, Dept Neurobiol, IL-76100 Rehovot, Israel

来源：

CHEMISTRY & BIOLOGY | 2012年 / 19卷 / 04期

基金：

美国国家卫生研究院;

关键词：

DIRECTED EVOLUTION; HYDROLYSIS; ENZYME; PON1; ENANTIOSELECTIVITY; STEREOISOMERS; PARAOXONASES; PROTECTION; INVERSION; TOXICITY;

D O I：

10.1016/j.chembiol.2012.01.017

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

070307 [化学生物学]; 071010 [生物化学与分子生物学];

摘要：

A preferred strategy for preventing nerve agents intoxication is catalytic scavenging by enzymes that hydrolyze them before they reach their targets. Using directed evolution, we simultaneously enhanced the activity of a previously described serum paraoxonase 1 (PON1) variant for hydrolysis of the toxic S-P isomers of the most threatening G-type nerve agents. The evolved variants show <= 340-fold increased rates and catalytic efficiencies of 0.2-5 x 10(7) M-1 min(-1). Our selection for prevention of acetylcholinesterase inhibition also resulted in the complete reversion of PON1's stereospecificity, from an enantiomeric ratio (E) < 6.3 x 10(-4) in favor of the R-P isomer of a cyclosarin analog in wild-type PON1, to E > 2,500 for the S-P isomer in an evolved variant. Given their ability to hydrolyze G-agents, these evolved variants may serve as broad-range G-agent prophylactics.

引用

页码：456 / 466

页数：11

共 42 条

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Directed evolution of mammalian paraoxonases PON1 and PON3 for bacterial expression and catalytic specialization [J].