title: Phosphorylation of the 20,000-Da myosin light chain isoforms of arterial smooth muscle by myosin light chain kinase and protein kinase C.
abstract: Phosphorylation of myosin light chain (LC) isoforms in arterial actomyosin can be induced by endogenous kinases upon addition of Mg2+ and ATP. The extent of phosphorylation in the presence of 2 mM ethylene glycol bis(beta-aminoethyl ether)N,N'-tetraacetic acid (EGTA), 0.2 mM Ca2+, or 0.2 mM Ca2+ plus calmodulin is 1.6, 2.1, and 2.3 mol phosphate/mol LC, respectively. Two-dimensional gel electrophoresis of actomyosin shows that the LC isoforms may be mono-, di-, and triphosphorylated. Tryptic phosphopeptide mapping of LC indicates that the radioactive phosphate is distributed to six peptides referred to as A through F. Phosphoamino acid analyses and the phosphopeptide maps of isolated LC, phosphorylated by either purified myosin light chain kinase or protein kinase C, reveal that myosin light chain kinase phosphorylates a serine residue in peptides A and B, and threonine plus serine residues in peptides C and D. Peptides E and F are phosphorylated by protein kinase C in serine and threonine residues, respectively. In actomyosin, with EGTA, phosphorylation of peptides E and F proceeds while phosphorylation of peptides A, B, C, and D is inhibited. Ca2+ and calmodulin enhance the phosphorylation of peptides A, B, C, and D, while phosphorylation of peptides E and F is decreased. In isolated LC, myosin light chain kinase preferentially phosphorylates the peptides A and B over C and D. Phosphorylation of peptides E and F in LC by protein kinase C promotes additional phosphorylation of peptides C and D by myosin light chain kinase, whereas phosphorylation of peptides A and B is diminished. The present data suggest that the phosphorylation of distinct sites in arterial myosin light chain by myosin light chain kinase and protein kinase C is interrelated.
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D001158:Arteries / Q000378:metabolism; D001665:Binding Sites; D066298:In Vitro Techniques; D008274:Magnesium / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D015879:Myosin Subfragments; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism*; D009218:Myosins / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D013552:Swine; D013912:Threonine / Q000378:metabolism
chemical_list: D015879:Myosin Subfragments; D010446:Peptide Fragments; D013912:Threonine; D012694:Serine; D000255:Adenosine Triphosphate; D011493:Protein Kinase C; D009219:Myosin-Light-Chain Kinase; D009218:Myosins; D008274:Magnesium