Conformational studies of myosin phosphorylated by protein kinase C.
Smooth muscle myosin from chicken gizzard is phosphorylated by Ca2+-activated phospholipid-dependent protein kinase, protein kinase C, as well as by Ca2+/calmodulin-dependent kinase, myosin light chain kinase (Endo, T., Naka, M., and Hidaka, H. (1982) Biochem. Biophys. Res. Commun. 105, 942-948). We have now demonstrated the effect of phosphorylation by protein kinase C on the smooth muscle myosin molecule. In glycerol/urea polyacrylamide gel electrophoresis the 20,000-dalton light chain phosphorylated by protein kinase C co-migrated with that phosphorylated by myosin light chain kinase. Moreover, the light chain phosphorylated by both kinases migrated more rapidly than did the light chain phosphorylated by either myosin light chain kinase or protein kinase C alone. Myosin phosphorylated by protein kinase C formed a bent 10 S monomer while that phosphorylated by myosin light chain kinase was an unfolded and extended 6 S monomer in the presence of 0.2 M KCl. In addition, myosin phosphorylated by kinases had a sedimentation velocity of 7.3 S, thereby suggesting that the myosin was partially unfolded. The unfolded myosin was visualized electron microscopically. The fraction in the looped form was higher when for myosin phosphorylated by both kinases higher than for that phosphorylated by light chain kinase alone. Therefore, phosphorylation by protein kinase C does not lead to the change in myosin conformation seen with myosin light chain kinase.
Umekawa
H
H
Naka
M
M
Inagaki
M
M
Onishi
H
H
Wakabayashi
T
T
Hidaka
H
H
D000818:Animals; D002645:Chickens; D004591:Electrophoresis, Polyacrylamide Gel; D005895:Gizzard, Avian / Q000032:analysis; D008854:Microscopy, Electron; D008970:Molecular Weight; D009130:Muscle, Smooth / Q000032:analysis; D009219:Myosin-Light-Chain Kinase; D009218:Myosins / Q000378:metabolism*; D010766:Phosphorylation; D011189:Potassium Chloride / Q000494:pharmacology; D011487:Protein Conformation; D011493:Protein Kinase C; D011494:Protein Kinases / Q000378:metabolism*
D011189:Potassium Chloride; D011494:Protein Kinases; D011493:Protein Kinase C; D009219:Myosin-Light-Chain Kinase; D009218:Myosins