title: Elucidation of calpain dependent phosphorylation of myosin light chain in human platelets.
abstract: Newly synthetized calpain inhibitors (CI-I approximately III) were used to prove potential participation of calpain in protein phosphorylation. CIs were about 1,000 times more potent against platelet calpain I than N-ethyl-maleimide (NEM) and an epoxy succinate derivative (E-64). CI-II inhibited 20K (myosin light chain) and 47K phosphorylation of Ca2+-stimulated lysed platelets as well as protein degradation (actin binding protein, P235). Both myosin light chain kinase (MLCK) and C-kinase dependent phosphorylation of 20K were inhibited by CI-II as demonstrated in phosphopeptides mapping. Electropermeabilized platelets (EP) were employed to examine the effects of CI-II on Ca2+ mediated reactions in non-lysed platelets. Phosphorylation of 20K and 47K induced by Ca2+ addition to EP was inhibited by CI-II, though secretory response was not modified. Only MLCK dependent phosphorylation of 20K was observed in Ca2+-activated EP, which was inhibited by CI-II. Collectively, the data indicated that calpain may activate both MLCK and C-kinase to phosphorylate 20K by partial proteolysis.
authors.affiliation: Second Department of Surgery, Osaka University Medical School, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Kajiwara
authors.forename: T
authors.initials: T
authors.lastname: Tsujinaka
authors.forename: M
authors.initials: M
authors.lastname: Sakon
authors.forename: J
authors.initials: J
authors.lastname: Kambayashi
authors.forename: T
authors.initials: T
authors.lastname: Ohshiro
authors.forename: T
authors.initials: T
authors.lastname: Murachi
authors.forename: T
authors.initials: T
authors.lastname: Mori
mesh_terms: D000167:Acriflavine; D001792:Blood Platelets / Q000378:metabolism*; D001798:Blood Proteins / Q000378:metabolism; D002154:Calpain / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D066298:In Vitro Techniques; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism; D009218:Myosins / Q000378:metabolism*; D010761:Phosphorus Radioisotopes; D010766:Phosphorylation; D014357:Trypsin
chemical_list: D001798:Blood Proteins; D010761:Phosphorus Radioisotopes; D000167:Acriflavine; D009219:Myosin-Light-Chain Kinase; D014357:Trypsin; D002154:Calpain; D009218:Myosins