Puntoni F, Villa-Moruzzi E
Department of Biomedicine, University of Pisa, Italy.
Mol Cell Biochem. 1997 Jun;171(1-2):115-20. doi: 10.1023/a:1006892103306.
Protein Phosphatase-1 is phosphorylated in vitro by cdc2-cyclin B (Villa-Moruzzi, FEBS Lett 304: 211-215, 1992). In the present study we show that all the three Phosphatase-1 isoforms, alpha, gamma 1, delta, are phosphorylated by cdc2-cyclin B. Phosphorylation is specific for this kinase and involves a C-terminal Thr. This site is most likely Thr 320 in alpha (shown by others to be phosphorylated also by cdc2-cyclin A). Such Thr is conserved in gamma 1, delta and in the testis-specific gamma 2, and is the only Thr that fits the cdc2-consensus sequence in the C-terminal region. Phosphorylation of Phosphatase-1 purified from skeletal muscle, which is a mixture of the alpha, gamma 1 and delta isoforms, is up to 0.4 mol/mol and induces 30-35% enzyme inactivation. Following tryptic proteolysis each isoform yields a distinct phosphopeptide map. This is in agreement with the different sequences of the isoforms in the C-terminal regions and may be useful to distinguish the isoforms in extracts from metabolically-labelled cells. Our results suggest that all the Phosphatase-1 isoforms may be potentially regulated at M-phase.
蛋白磷酸酶-1在体外被细胞周期蛋白依赖性激酶2-细胞周期蛋白B磷酸化（Villa-Moruzzi，《欧洲分子生物学学会联合会快报》304：211-215，1992年）。在本研究中，我们发现蛋白磷酸酶-1的所有三种同工型，即α、γ1、δ，均被细胞周期蛋白依赖性激酶2-细胞周期蛋白B磷酸化。这种磷酸化作用对该激酶具有特异性，且涉及一个C末端苏氨酸。此位点很可能是α中的苏氨酸320（其他人也表明它也可被细胞周期蛋白依赖性激酶2-细胞周期蛋白A磷酸化）。这样的苏氨酸在γ1、δ以及睾丸特异性的γ2中是保守的，并且是C末端区域中唯一符合细胞周期蛋白依赖性激酶2一致序列的苏氨酸。从骨骼肌中纯化的蛋白磷酸酶-1是α、γ1和δ同工型的混合物，其磷酸化程度高达0.4摩尔/摩尔，并导致30 - 35%的酶失活。经胰蛋白酶消化后，每种同工型产生独特的磷酸肽图谱。这与C末端区域同工型的不同序列一致，并且可能有助于区分来自代谢标记细胞提取物中的同工型。我们的结果表明，所有蛋白磷酸酶-1同工型在M期可能都受到潜在调控。
