解脂耶氏酵母中一种不依赖cAMP的Ca2(+)抑制型鱼精蛋白激酶的特性分析。

Characterization of a cAMP-independent Ca2(+)-inhibited protamine kinase from Candida lipolytica.

作者信息

Rahmatullah M, Brenner D L, Wooten M W, Weete J D

机构信息

Department of Botany and Microbiology, Auburn University, AL 36849.

出版信息

Biochem Biophys Res Commun. 1991 Mar 15;175(2):500-6. doi: 10.1016/0006-291x(91)91592-z.

DOI:10.1016/0006-291x(91)91592-z

PMID:1850244

Abstract

A cAMP-independent protamine kinase has been purified from extracts of the yeast Candida lipolytica by ion-exchange and affinity chromatography. Two subunits with apparent Mr's of 52,000 and 36,000 were resolved by SDS-PAGE. The purified kinase exhibited about 20% activity with casein and histone Type VII-S as substrates relative to protamine. The enzyme was inactive against other protein substrates tested, and was essentially insensitive to AMP, cAMP, cGMP up to 0.2 mM, the polyamines spermine and spermidine up to 1 mM, N-ethylmaleimide (5 mM), 2-mercaptoethanol (20 mM), or dithiothreitol (2 mM), and several cations like Zn2+, N1+, or Co2+ at 0.1 mM each. Ca2+ at 3 mM inhibited protamine kinase activity by 50%, which was reversed by EGTA.

摘要

一种不依赖环磷酸腺苷（cAMP）的鱼精蛋白激酶已通过离子交换和亲和层析从解脂假丝酵母提取物中纯化出来。通过十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳（SDS - PAGE）分离出了两个表观分子量分别为52,000和36,000的亚基。相对于鱼精蛋白，纯化后的激酶以酪蛋白和VII - S型组蛋白为底物时表现出约20%的活性。该酶对所测试的其他蛋白质底物无活性，并且对高达0.2 mM的AMP、cAMP、cGMP、高达1 mM的多胺精胺和亚精胺、5 mM的N - 乙基马来酰亚胺、20 mM的2 - 巯基乙醇或2 mM的二硫苏糖醇以及每种浓度为0.1 mM的几种阳离子如Zn2 +、Ni +或Co2 +基本不敏感。3 mM的Ca2 +可抑制鱼精蛋白激酶活性50%，而乙二醇双乙醚二胺四乙酸（EGTA）可逆转这种抑制作用。