Ansai T, Dupuy L C, Barik S
Department of Biochemistry and Molecular Biology, MSB2140, University of South Alabama College of Medicine, Mobile, Alabama 36688-0002, USA.
J Biol Chem. 1996 Oct 4;271(40):24401-7. doi: 10.1074/jbc.271.40.24401.
The protein phosphatase encoded by coliphage lambda (PPlambda) was found to be the equivalent of the minimal catalytic core of serine/threonine protein phosphatases (PP) by biochemical and mutational criteria. Bacterially expressed truncated versions of PP1 and PP5 phosphatases, representing the catalytic cores homologous to PPlambda, exhibited potent phosphatase activity. Unlike full-length PP1, but like PPlambda, the recombinant cores could use casein, p-nitrophenyl phosphate, and a wide variety of peptides as substrates and were resistant to okadaic acid, microcystin-LR, and trypsin. Mutations of His173, Asp208, or Arg221 had little effect on the activity of the PP1 core protein, indicating its closer identity with PPlambda than with full-length PP1. Terminal deletions of a few amino acids of the cores destroyed their activity, supporting their minimal nature. Analysis of PPlambda mutants suggested an influence of the substrate on metal ion binding. The minimal length of a phosphopeptide substrate of PPlambda appeared to be a phosphorylated serine/threonine flanked by 1 or 2 amino acid residues on either side, the N-terminal ones being more effective.
通过生化和突变标准发现，噬菌体λ编码的蛋白磷酸酶（PPlambda）等同于丝氨酸/苏氨酸蛋白磷酸酶（PP）的最小催化核心。在细菌中表达的PP1和PP5磷酸酶的截短版本，代表与PPlambda同源的催化核心，表现出强大的磷酸酶活性。与全长PP1不同，但与PPlambda一样，重组核心可以使用酪蛋白、对硝基苯磷酸酯和多种肽作为底物，并且对冈田酸、微囊藻毒素-LR和胰蛋白酶具有抗性。His173、Asp208或Arg221的突变对PP1核心蛋白的活性影响很小，表明其与PPlambda的亲缘关系比与全长PP1更近。核心区域末端缺失几个氨基酸会破坏其活性，支持了它们的最小性质。对PPlambda突变体的分析表明底物对金属离子结合有影响。PPlambda磷酸肽底物的最小长度似乎是一个磷酸化的丝氨酸/苏氨酸，两侧各有1或2个氨基酸残基，N端的氨基酸残基更有效。
