Stereospecificity in the enzymatic hydrolysis of cyclosarin (GF)
Stereospecificity in the enzymatic hydrolysis of cyclosarin (GF)
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DOI：
10.1016/j.enzmictec.2005.04.004
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发表时间：
2005-10-03
影响因子：
3.4
通讯作者：
Hill, C
中科院分区：
文献类型：
--
作者：
Harvey, SP;Kolakowski, JE;Hill, C
酶催化是加速G型有机磷神经毒剂水解率的方法之一。本文研究了几种酶催化环沙林(GF，O-环己基甲基氟膦)水解的立体专一性。杂交性在3 mm纤维上不明显，但在0.5 mm纤维上明显。这种差异明显是由于氟化物催化底物外消旋所致。Alteromonas sp.通过对剩余未反应的(-)GF异构体的GC分析，发现JD6.5有机磷酸失水解酶(OPAA)、盐生假单胞菌OPAA和野生型磷酸三酯酶(PTE)都优先催化(+)GF异构体的水解。乙酰胆碱酯酶抑制实验表明，纯化的(-)GF异构体的毒性大约是外消旋混合物的两倍。一个Pte突变体H254G/H259W/L303T逆转了天然Pte的立体特异性，并优先催化(-)GF异构体的水解，这一点从它与Alteromonas sp.的互补中可以看出。JD6.5 OPAA，对剩余的(+)GF异构体进行GC分析。这一程序还允许通过酶降解单独制备两种GF异构体中的任何一种，然后提取剩余的异构体。(C)2005 Elsevier Inc.保留所有权利。
Enzymatic catalysis is one means of accelerating the rate of hydrolysis of G-type organophosphorus nerve agents. Here, the stereospecificity of the catalysis of cyclosarin (GF, O-cyclohexyl methylphosphonofluoridate) hydrolysis by several enzymes was investigated. Stercospecificity was not evident at 3 mM GF but was evident at 0.5 mM GF. The differential effect was apparently due to fluoride-catalyzed racemization of the substrate. Alteromonas sp. JD6.5 organophosphorus acid anhydrolase (OPAA), Alteromonas haloplanktis OPAA and the wild-type phosphotriesterase (PTE) enzymes were all found to catalyze preferentially the hydrolysis of the (+)GF isomer, as determined by GC analysis of the remaining unreacted (-)GF isomer. Acetylcholinesterase inhibition experiments showed the purified (-)GF isomer to be approximately twice as toxic as the racemic mixture. One PTE mutant, H254G/H259W/L303T, was found to reverse the native PTE stereospecificity and preferentially catalyze the hydrolysis of the (-)GF isomer, as shown by its complementation of Alteromonas sp. JD6.5 OPAA and by GC analysis of the remaining (+)GF isomer. This procedure also permitted the individual preparation of either of the two GF isomers by enzymatic degradation followed by extraction of the remaining isomer. (c) 2005 Elsevier Inc. All rights reserved.