The 204-kDa smooth muscle myosin heavy chain is phosphorylated in intact cells by casein kinase II on a serine near the carboxyl terminus.
The heavy chain of smooth muscle myosin was found to be phosphorylated following immunoprecipitation from cultured bovine aortic smooth muscle cells. Of a variety of serine/threonine kinases assayed, only casein kinase II and calcium/calmodulin-dependent protein kinase II phosphorylated the smooth muscle myosin heavy chain to a significant extent in vitro. Two-dimensional maps of tryptic peptides derived from heavy chains phosphorylated in cultured cells revealed one major and one minor phosphopeptide. Identical tryptic peptide maps were obtained from heavy chains phosphorylated in vitro with casein kinase II but not with calcium/calmodulin-dependent protein kinase II. Of note, the 204-kDa smooth muscle myosin heavy chain but not the 200-kDa heavy chain isoform was phosphorylated by casein kinase II. Partial sequence of the tryptic phosphopeptides generated following phosphorylation by casein kinase II yielded Val-Ile-Glu-Asn-Ala-Asp-Gly-Ser*-Glu-Glu-Glu-Val. The Ser* represents the Ser(PO4) which is in an acidic environment, as is typical for casein kinase II phosphorylation sites. By comparison with the deduced amino acid sequence for rabbit uterine smooth muscle myosin (Nagai, R., Kuro-o, M., Babij, P., and Periasamy, M. (1989) J. Biol. Chem. 264, 9734-9737), we have localized the phosphorylated serine residue to the non-helical tail of the 204-kDa isoform of the smooth muscle myosin heavy chain. The ability of the 204-kDa isoform, but not the 200-kDa isoform, to serve as a substrate for casein kinase II suggests that these two isoforms can be regulated differentially.
Kelley
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CA
Laboratory of Molecular Cardiology, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892.
Adelstein
R S
RS
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000302:isolation & purification; D000818:Animals; D001011:Aorta / Q000378:metabolism*; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D047388:Casein Kinases; D002417:Cattle; D015180:Electrophoresis, Gel, Two-Dimensional; D005260:Female; D006801:Humans; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009130:Muscle, Smooth / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D009218:Myosins / Q000302:isolation & purification / Q000378:metabolism*; D010748:Phosphopeptides / Q000302:isolation & purification; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D011817:Rabbits; D012694:Serine; D014599:Uterus / Q000378:metabolism*
D000596:Amino Acids; D046911:Macromolecular Substances; D010748:Phosphopeptides; D012694:Serine; D011494:Protein Kinases; D047388:Casein Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D009218:Myosins