Phosphorylation of the large subunit of myosin phosphatase and inhibition of phosphatase activity.
The partially purified myosin-bound phosphatase had an associated protein kinase that phosphorylated the holoenzyme, primarily on the large (130-kDa) subunit. Phosphorylation of the 130-kDa subunit resulted in inhibition of phosphatase activity. The major site of phosphorylation was threonine 654 of the 130-kDa subunit or threonine 695 of the 133-kDa isoform. Phosphorylation of the large subunit did not dissociate the holoenzyme. Dephosphorylation of the large subunit was achieved by the holoenzyme, and addition of the catalytic subunit of the type 2A enzyme did not increase the rate of dephosphorylation. The associated kinase was inhibited by chelerythrine, with half-maximal inhibition at approximately 5 microM (in 150 microM ATP). The associated kinase phosphorylated two synthetic peptides, one corresponding to the sequence flanking the phosphorylated threonine, i.e. 648-661 of the 130-kDa subunit, and the other to a known protein kinase C substrate, i.e. a modified sequence from the autoinhibitory region of epsilon protein kinase C. The associated kinase was activated by arachidonic and oleic acid and to a lesser extent by myristic acid. The protein kinase that phosphorylated the 130-kDa subunit and resulted in inhibition of myosin phosphatase activity was not identified.
Ichikawa
K
K
Muscle Biology Group, University of Arizona, Tucson, Arizona 85721, USA.
Ito
M
M
Hartshorne
D J
DJ
D000255:Adenosine Triphosphate / Q000378:metabolism; D000470:Alkaloids; D000595:Amino Acid Sequence; D000818:Animals; D001345:Autoradiography; D053119:Benzophenanthridines; D002645:Chickens; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005230:Fatty Acids, Nonesterified / Q000494:pharmacology; D005895:Gizzard, Avian / Q000201:enzymology; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009130:Muscle, Smooth / Q000201:enzymology*; D043263:Myosin-Light-Chain Phosphatase; D010617:Phenanthridines / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D010750:Phosphoproteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D010761:Phosphorus Radioisotopes; D010766:Phosphorylation; D014422:Turkeys
D000470:Alkaloids; D053119:Benzophenanthridines; D004791:Enzyme Inhibitors; D005230:Fatty Acids, Nonesterified; D046911:Macromolecular Substances; D010617:Phenanthridines; D010750:Phosphoproteins; D010761:Phosphorus Radioisotopes; D000255:Adenosine Triphosphate; C016299:chelerythrine; D010749:Phosphoprotein Phosphatases; D043263:Myosin-Light-Chain Phosphatase