Lombardo D, Guy O
Biochim Biophys Acta. 1981 Feb 13;657(2):425-37. doi: 10.1016/0005-2744(81)90328-4.
Transfer reactions catalyzed by human pancreatic carboxylic-ester hydrolase (EC 3.1.1.1) were studied in the presence of methanol and butanol as nucleophiles. The addition of alcohols produced an increase in the total rate of 4-nitrophenyl acetate and n-propylthiol acetate disappearance and a concomitant slow decrease of the hydrolysis rate. These results indicate a competitive partitioning of an acyl-enzyme intermediate between water and nucleophile. Moreover, a strong inhibition of the rates of hydrolysis of methyl butyrate and triacetin by nucleophiles is in agreement with a rate-limiting acylation step. The kinetic data and a trans-ester characterization argue in favor of the formation of an acyl-enzyme intermediate and a two-step reaction mechanism, acylation and deacylation both being rate-limiting. The experiments performed with 4-nitrophenyl acetate show the existence of a nucleophile binding site.
在以甲醇和丁醇作为亲核试剂的情况下，对人胰腺羧酸酯水解酶（EC 3.1.1.1）催化的转移反应进行了研究。添加醇类导致乙酸对硝基苯酯和乙酸正丙硫醇的总消失速率增加，同时水解速率缓慢下降。这些结果表明酰基酶中间体在水和亲核试剂之间存在竞争性分配。此外，亲核试剂对丁酸甲酯和三醋精水解速率的强烈抑制与限速酰化步骤一致。动力学数据和转酯表征支持酰基酶中间体的形成以及两步反应机制，酰化和脱酰化均为限速步骤。用乙酸对硝基苯酯进行的实验表明存在亲核试剂结合位点。
