Department of Chemistry, Texas A&M University, College Station, Texas 77843, United States.
US Army DEVCOM-CBC, FCDD-CBR-CC E3400, 5183 Blackhawk Rd. Aberdeen Proving Ground, Aberdeen, Maryland 21010, United States.
Biochemistry. 2021 Sep 28;60(38):2875-2887. doi: 10.1021/acs.biochem.1c00361. Epub 2021 Sep 8.
The G-type nerve agents, sarin (GB), soman (GD), and cyclosarin (GF), are among the most toxic compounds known. Much progress has been made in evolving the enzyme phosphotriesterase (PTE) from for the decontamination of the G-agents; however, the extreme toxicity of the G-agents makes the use of substrate analogues necessary. Typical analogues utilize a chromogenic leaving group to facilitate high-throughput screening, and substitution of an -methyl for the -methyl group found in the G-agents, in an effort to reduce toxicity. Till date, there has been no systematic evaluation of the effects of these substitutions on catalytic activity, and the presumed reduction in toxicity has not been tested. A series of 21 G-agent analogues, including all combinations of -methyl, -nitrophenyl, and thiophosphate substitutions, have been synthesized and evaluated for their ability to unveil the stereoselectivity and catalytic activity of PTE variants against the authentic G-type nerve agents. The potential toxicity of these analogues was evaluated by measuring the rate of inactivation of acetylcholinesterase (AChE). All of the substitutions reduced inactivation of AChE by more than 100-fold, with the most effective being the thiophosphate analogues, which reduced the rate of inactivation by about 4-5 orders of magnitude. The analogues were found to reliably predict changes in catalytic activity and stereoselectivity of the PTE variants and led to the identification of the BHR-30 variant, which has no apparent stereoselectivity against GD and a / of 1.4 × 10, making it the most efficient enzyme for GD decontamination reported till date.
G 型神经毒剂，沙林（GB）、梭曼（GD）和环沙林（GF），是已知毒性最强的化合物之一。在开发酶膦酸三酯酶（PTE）用于 G 型毒剂解毒方面已经取得了很大进展；然而，G 型毒剂的极高毒性使得必须使用底物类似物。典型的类似物利用生色离去基团来促进高通量筛选，并用 -甲基取代 G 型毒剂中的 -甲基，以努力降低毒性。迄今为止，尚未对这些取代基对催化活性的影响进行系统评估，而且假定的毒性降低也未经过测试。已经合成了一系列 21 种 G 型毒剂类似物，包括 -甲基、-硝基苯基和硫代膦酸酯取代物的所有组合，并评估了它们对 PTE 变体对真实 G 型神经毒剂的立体选择性和催化活性的揭示能力。通过测量乙酰胆碱酯酶（AChE）的失活率来评估这些类似物的潜在毒性。所有取代基都使 AChE 的失活率降低了 100 多倍，最有效的是硫代膦酸类似物，其失活率降低了约 4-5 个数量级。发现这些类似物能够可靠地预测 PTE 变体的催化活性和立体选择性的变化，并导致鉴定出 BHR-30 变体，该变体对 GD 没有明显的立体选择性，/为 1.4×10，使其成为迄今为止报道的 GD 解毒最有效的酶。
