3-溴丙酮酸对大肠杆菌异柠檬酸裂解酶的烷基化作用。

Alkylation of isocitrate lyase from Escherichia coli by 3-bromopyruvate.

作者信息

Ko Y H, McFadden B A

机构信息

Biochemistry/Biophysics Program, Washington State University, Pullman 99164-4660.

出版信息

Arch Biochem Biophys. 1990 May 1;278(2):373-80. doi: 10.1016/0003-9861(90)90273-2.

DOI:10.1016/0003-9861(90)90273-2

PMID:2183722

Abstract

The inactivation of tetrameric isocitrate lyase from Escherichia coli by 3-bromopyruvate, exhibiting saturation kinetics, is accompanied by the loss of one sulfhydryl per subunit. The substrates glyoxylate and isocitrate protect against inactivation whereas the substrate succinate does not. The modification by 3-bromopyruvate (equimolar to subunits) imparts striking resistance to digestion of isocitrate lyase by trypsin, chymotrypsin, and V8 protease as well as a major decrease in the intensity of tryptophan fluorescence. After alkylation, the sequence Gly-His-Met-Gly-Gly-Lys is found following the modified Cys residue in the tryptic peptide representing positions 196-201. Thus Cys195 is alkylated by 3-bromopyruvate.

摘要

3-溴丙酮酸使来自大肠杆菌的四聚体异柠檬酸裂解酶失活，呈现出饱和动力学，且每个亚基会损失一个巯基。底物乙醛酸和异柠檬酸可防止酶失活，而底物琥珀酸则不能。3-溴丙酮酸（与亚基等摩尔）的修饰使异柠檬酸裂解酶对胰蛋白酶、糜蛋白酶和V8蛋白酶的消化具有显著抗性，同时色氨酸荧光强度大幅降低。烷基化后，在代表196 - 201位的胰蛋白酶肽段中，修饰的半胱氨酸残基之后发现了Gly-His-Met-Gly-Gly-Lys序列。因此，Cys195被3-溴丙酮酸烷基化。