非洲爪蟾卵母细胞中多阳离子刺激的蛋白磷酸酶底物特异性的调节

Modulation of the substrate specificity of the polycation-stimulated protein phosphatase from Xenopus laevis oocytes.

作者信息

Hermann J, Cayla X, Dumortier K, Goris J, Ozon R, Merlevede W

机构信息

Afdeling Biochemie, Faculteit Geneeskunde, Katholieke Universiteit Leuven, Belgium.

出版信息

Eur J Biochem. 1988 Apr 5;173(1):17-25. doi: 10.1111/j.1432-1033.1988.tb13961.x.

DOI:10.1111/j.1432-1033.1988.tb13961.x

PMID:2833390

Abstract

A polycation-stimulated (PCS) protein phosphatase was isolated in high yield (280 micrograms/100 g ovaries) from Xenopus laevis oocytes through a procedure involving a tyrosine-agarose hydrophobic chromatography. The 220-kDa enzyme contains a 35-kDa and a 62-kDa subunit. It was identified as the low-Mr polycation-stimulated (PCSL) protein phosphatase. The labile p-nitrophenyl phosphatase activity, copurifying with the phosphorylase phosphatase activity, can be increased severalfold by preincubating the purified enzyme with ATP, its analogues or PPi. This activation is time-dependent and accompanied by a parallel decrease of the phosphorylase phosphatase activity. Although the stimulation was antagonized by metal ions during the preincubation, the basal and ATP-stimulated p-nitrophenyl phosphatase requires Mg2+ or Mn2+ in the assay, with pH optima of 8.5-9 and 7.5 respectively.

摘要

通过一种涉及酪氨酸 - 琼脂糖疏水色谱的方法，从非洲爪蟾卵母细胞中高产率地分离出一种多阳离子刺激（PCS）蛋白磷酸酶（280微克/100克卵巢）。这种220 kDa的酶含有一个35 kDa和一个62 kDa的亚基。它被鉴定为低分子量多阳离子刺激（PCSL）蛋白磷酸酶。与磷酸化酶磷酸酶活性共纯化的不稳定对硝基苯磷酸酶活性，可通过将纯化的酶与ATP、其类似物或焦磷酸预孵育而提高数倍。这种激活是时间依赖性的，同时伴随着磷酸化酶磷酸酶活性的平行降低。尽管在预孵育过程中金属离子会拮抗这种刺激，但基础的和ATP刺激的对硝基苯磷酸酶在测定中需要Mg2+或Mn2+，pH最佳值分别为8.5 - 9和7.5。