Phosphorylation of myorod (catchin) by kinases tightly associated to molluscan and vertebrate smooth muscle myosins.
Myorod, also known as catchin, a newly discovered component of molluscan smooth muscle thick filaments, is an alternative product of the myosin heavy chain gene. It contains a C-terminal rod part that is identical to that part of myosin and a unique N-terminal domain that is very small relative to the myosin head domain. The role of myorod in contraction or relaxation of this muscle type is unknown. In the present study we demonstrated that myorod was phosphorylated not only by a kinase endogenous to molluscan myosin and twitchin but also to vertebrate smooth muscle myosin light chain kinase (MLCK). The rates and maximal levels of phosphorylation were up to threefold higher than those observed by protein kinase A with clear optima at the physiological salt concentrations. Using a mild digestion with chymotrypsin we isolated an 11 kDa phosphopeptide and showed that the phosphorylation site was located at the N-terminal domain of myorod at Thr 141 position. The sequence around this site exhibited a high degree of similarity to that expected for the substrate recognition site of MLCK. The phosphorylation rates strongly depended on the ionic conditions indicating that this site could be readily sterically blocked during myorod polymerization. Another component of the thick filaments involved in regulation of the catch state, twitchin, was phosphorylated by MLCK and exhibited endogenous myorod kinase and MLCK activities. A possible role of these phosphorylation reactions in the regulation of molluscan smooth muscles is discussed.
Sobieszek
Apolinary
A
Institute for Biomedical Aging Research, Life Science Center, Austrian Academy of Sciences, Mitterweg 24, A-6020 Innsbruck, Austria. ASobieszek@oeaw.ac.at
Matusovsky
Oleg S
OS
Permyakova
Tatyana V
TV
Sarg
Bettina
B
Lindner
Herbert
H
Shelud'ko
Nikolay S
NS
D000818:Animals; D001665:Binding Sites; D049872:Bivalvia / Q000378:metabolism*; D002918:Chymotrypsin / Q000737:chemistry; D017868:Cyclic AMP-Dependent Protein Kinases / Q000737:chemistry; D009124:Muscle Proteins / Q000737:chemistry* / Q000302:isolation & purification; D009130:Muscle, Smooth / Q000378:metabolism*; D018995:Myosin Heavy Chains / Q000737:chemistry*; D009219:Myosin-Light-Chain Kinase / Q000737:chemistry*; D010748:Phosphopeptides / Q000737:chemistry; D010766:Phosphorylation; D014714:Vertebrates
D009124:Muscle Proteins; D010748:Phosphopeptides; D017868:Cyclic AMP-Dependent Protein Kinases; D009219:Myosin-Light-Chain Kinase; D002918:Chymotrypsin; D018995:Myosin Heavy Chains