Purification and characterization of a novel protein phosphatase highly specific for ribosomal protein S6.
Purification and characterization of a novel protein phosphatase highly specific for ribosomal protein S6.
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发表时间：
1989-01
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通讯作者：
J. Andres;J. Maller
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作者：
J. Andres;J. Maller
核糖体蛋白S6是真核核糖体的主要磷蛋白，在多种有丝分裂刺激下，核糖体在丝氨酸残基上发生多重磷酸化。本文研究了非洲爪哇卵巢和卵中能使S6脱磷的主要蛋白磷酸酶。两种称为峰I和峰II的酶解释了卵母细胞和卵子中S6磷酸酶的大部分活性。经凝胶过滤，峰I酶的表观Mr为200,000，使磷酸化酶激酶和磷酸化酶a的β亚基去磷酸化，并被抑制物1和抑制物2抑制，表明其与蛋白磷酸酶1相似。经甘油梯度离心法，峰II酶纯化了12,000多倍，表观MR=55,000。这种磷酸酶可以去磷酸化S6中的所有位点，但不能去磷酸化磷酸化酶a或磷酸化酶激酶。但抑制物1和抑制物2的纳摩尔浓度对其有抑制作用。这些结果表明，峰值II酶代表了一类新的高度特异的蛋白磷酸酶，并提示抑制物1和抑制物2抑制细胞提取物中的去磷酸化不能作为蛋白磷酸酶1参与细胞过程的充分标准。
Ribosomal protein S6 is the principal phosphoprotein of the eucaryotic ribosome that becomes multiply phosphorylated on serine residues in response to a wide variety of mitogenic stimuli. In this paper the principal protein phosphatases able to dephosphorylate S6 were characterized in Xenopus laevis ovary and eggs. Two enzymes termed peak I and peak II were found to account for most S6 phosphatase activity in both oocytes and eggs. The peak I enzyme had an apparent Mr of 200,000 on gel filtration, dephosphorylated the beta subunit of phosphorylase kinase and phosphorylase a, and was inhibited by inhibitor 1 and inhibitor 2, suggesting it was similar to protein phosphatase 1. The peak II enzyme was purified over 12,000-fold and had an apparent Mr = 55,000 on glycerol gradient centrifugation. This phosphatase could dephosphorylate all sites in S6 but was unable to dephosphorylate phosphorylase a or phosphorylase kinase. However, it was inhibited by nanomolar concentrations of inhibitor 1 and inhibitor 2. These results indicate the peak II enzyme represents a new class of highly specific protein phosphatase and suggest that inhibition of dephosphorylation in cellular extracts by inhibitor 1 and inhibitor 2 is not a sufficient criterion for implicating protein phosphatase 1 in a cellular process.