期刊
JOURNAL OF BIOLOGICAL CHEMISTRY
卷 276, 期 43, 页码 39858-39863出版社
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M107302200
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资金
- NCI NIH HHS [CA 40042] Funding Source: Medline
- NIGMS NIH HHS [GM 56362] Funding Source: Medline
Contractility of smooth muscle and non-muscle micro. filaments involves phosphorylation of myosin II light chain. Myosin light chain phosphatase (MLCP) is specifically inhibited by the protein kinase C-potentiated inhibitor protein of 17 kDa, called CPI-17, as part of Ca2+ sensitization of vascular smooth muscle contraction. Phosphorylation of Thr(38) in CPI-17 enhances inhibitory potency toward MLCP over 1000-fold. In this study we mapped regions of CPI-17 required for inhibition and investigated the mechanism using deletion and point mutants. Deletion of either the N-terminal 34 residues or C-terminal 27 residues gave no change in the IC50 of either phospho- or unphospho-CPI-17. However, further deletion to give CPI-17 proteins of 1-102,1-89,1-76, and 1-67, resulted in much higher IC50 values. The results indicate there is a minimal inhibitory domain between residues 35 and 120. A single Ala substitution at Tyr 41 eliminated phosphorylation-dependent inhibition, and phospho-Thr(38) in the Y41A protein was efficiently dephosphorylated by MLCP itself. The wild type CPI-17 expressed in fibroblast-induced bundling and contraction of actomyosin filaments, whereas expression of the Y41A protein had no obvious effects. Thus, a central domain of CPI-17(35-120) including phospho-Thr(38) is necessary for recognition by myosin phosphatase and Tyr(41) arrests dephosphorylation, thereby producing inhibition.
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