核糖核酸酶F1的活性位点谷氨酸残基被碘乙酸羧甲基化。

Carboxymethylation of an active site glutamic acid residue of ribonuclease F1 iodoacetate.

作者信息

Yoshida H, Hanazawa H

机构信息

Department of Chemistry, Faculty of Science, Tohoku University, Sendai, Japan.

出版信息

Biochimie. 1989 May;71(5):687-92. doi: 10.1016/0300-9084(89)90163-6.

DOI:10.1016/0300-9084(89)90163-6

PMID:2569896

Abstract

Ribonuclease (RNase) F1 was inactivated by incubation with an excess amount of iodoacetate at pH 5.5, 37 degrees C according to pseudo first-order kinetics. It was protected to various degrees, from inactivation by nucleotides, among which guanosine 2'-phosphate was most effective. The pseudo first-order rate constant was proportional to the reagent concentration, indicating that the reaction in reality follows second-order kinetics. The second-order rate constant was determined to be 25 x 10(-4) M-1 s-1. The inactivation rate was maximal at pH 5.5-6.0. When iodo[2-14C]acetate was used as the reagent, the stoichiometry of incorporation was determined to be 1.1 mol carboxymethyl group per mol of RNase F1 and glutamic acid residue 58 was assigned as the site of modification.

摘要

根据准一级动力学，在pH 5.5、37℃下，核糖核酸酶（RNase）F1与过量碘乙酸孵育后失活。它受到核苷酸不同程度的保护而不被失活，其中2'-磷酸鸟苷最为有效。准一级速率常数与试剂浓度成正比，表明该反应实际上遵循二级动力学。二级速率常数测定为25×10⁻⁴ M⁻¹ s⁻¹。失活速率在pH 5.5 - 6.0时最大。当使用碘[2-¹⁴C]乙酸作为试剂时，确定每摩尔RNase F1的羧甲基掺入化学计量比为1.1摩尔，并且将谷氨酸残基58指定为修饰位点。