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Stereoselective Formation of Multiple Reaction Products by the Phosphotriesterase from Sphingobium sp. TCM1.
Biochemistry ( IF 2.9 ) Pub Date : 2020-03-17 , DOI: 10.1021/acs.biochem.0c00089 Andrew N Bigley 1 , Tamari Narindoshvili 1 , Dao Feng Xiang 1 , Frank M Raushel 1, 2
Biochemistry ( IF 2.9 ) Pub Date : 2020-03-17 , DOI: 10.1021/acs.biochem.0c00089 Andrew N Bigley 1 , Tamari Narindoshvili 1 , Dao Feng Xiang 1 , Frank M Raushel 1, 2
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Organophosphate flame retardants are used to inhibit combustion and increase plasticity in plastics and durable foams. While not neurotoxic, these compounds are potential carcinogens, endocrine disrupters, and developmental toxins. The phosphotriesterase from Sphingobium sp. TCM1 (Sb-PTE) is unique among phosphotriesterase enzymes for its ability to hydrolyze these compounds and its ability to hydrolyze any one of the three different ester bonds within a given substrate. In some cases, the extent of hydrolysis of a methyl ester exceeds that of a p-nitrophenyl ester within a single substrate. There is a stereochemical component to this hydrolysis where the two enantiomers of chiral substrates give different product ratios. To investigate the stereoselectivity for the product distribution of Sb-PTE, a series of 24 phosphotriesters were synthesized with all possible combinations of methyl, cyclohexyl, phenyl, and p-nitrophenyl esters. Prochiral compounds were made chiral by differential isotopic labeling using a chemo/enzymatic strategy, which allowed the differentiation of hydrolysis for each ester in all but two compounds. The rate equations for this unique enzymatic mechanism were derived; the product ratios were determined for each substrate, and the individual kinetic constants for hydrolysis of each ester within each substrate were measured. The findings are consistent with the rate-limiting step for substrate hydrolysis catalyzed by Sb-PTE being the formation of a phosphorane-like intermediate and the kinetic constants and product ratios being dictated by a combination of transition state energies, inductive effects, and stereochemical constraints.
中文翻译：
鞘氨醇单胞菌的磷酸三酯酶立体反应形成多个反应产物。中医1。
有机磷酸酯阻燃剂用于抑制燃烧并提高塑料和耐用泡沫塑料的可塑性。尽管没有神经毒性，但这些化合物是潜在的致癌物，内分泌干扰物和发育毒素。鞘氨醇单胞菌属的磷酸三酯酶。TCM1（Sb -PTE）在磷酸三酯酶中是独一无二的，因为它具有水解这些化合物的能力以及其水解给定底物中三个不同酯键中任何一个的能力。在一些情况下，甲酯的水解程度超过一个的p在单个底物中的-硝基苯基酯。该水解具有立体化学成分，其中手性底物的两种对映异构体给出不同的产物比率。为了研究Sb- PTE产物分布的立体选择性，合成了一系列24种磷酸三酯，其中甲基，环己基，苯基和p的所有可能组合-硝基苯酯。使用化学/酶促策略通过差异同位素标记，将手性化合物制成手性化合物，从而可以区分除两种化合物以外的所有酯的水解。推导了这种独特的酶促机理的速率方程。确定每种底物的产物比率，并测量每种底物中每种酯的水解的动力学常数。这些发现与Sb- PTE催化的底物水解的限速步骤一致，后者是磷烷样中间体的形成，动力学常数和产物比率是由过渡态能量，感应效应和立体化学约束条件共同决定的。 。
更新日期：2020-03-19
中文翻译：
鞘氨醇单胞菌的磷酸三酯酶立体反应形成多个反应产物。中医1。
有机磷酸酯阻燃剂用于抑制燃烧并提高塑料和耐用泡沫塑料的可塑性。尽管没有神经毒性，但这些化合物是潜在的致癌物，内分泌干扰物和发育毒素。鞘氨醇单胞菌属的磷酸三酯酶。TCM1（Sb -PTE）在磷酸三酯酶中是独一无二的，因为它具有水解这些化合物的能力以及其水解给定底物中三个不同酯键中任何一个的能力。在一些情况下，甲酯的水解程度超过一个的p在单个底物中的-硝基苯基酯。该水解具有立体化学成分，其中手性底物的两种对映异构体给出不同的产物比率。为了研究Sb- PTE产物分布的立体选择性，合成了一系列24种磷酸三酯，其中甲基，环己基，苯基和p的所有可能组合-硝基苯酯。使用化学/酶促策略通过差异同位素标记，将手性化合物制成手性化合物，从而可以区分除两种化合物以外的所有酯的水解。推导了这种独特的酶促机理的速率方程。确定每种底物的产物比率，并测量每种底物中每种酯的水解的动力学常数。这些发现与Sb- PTE催化的底物水解的限速步骤一致，后者是磷烷样中间体的形成，动力学常数和产物比率是由过渡态能量，感应效应和立体化学约束条件共同决定的。 。
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