A phosphorylated light-chain component of myosin from skeletal muscle.
1. The low-molecular-weight components of myosin from rabbit skeletal muscle migrated as four bands on polyacrylamide-gel electrophoresis in 8m-urea but only as three in systems containing sodium dodecyl sulphate. The two bands of intermediate mobility in 8m-urea (Ml(2) and Ml(3)) had identical mobilities in sodium dodecyl sulphate. 2. The isolation of pure samples of all four low-molecular-weight components by DEAE-Sephadex chromatography is described. 3. The amino acid compositions of components Ml(2) and Ml(3) were identical. Further analyses showed the presence of 1 mol of phosphate/18500g of component Ml(2) and less than 10% of this amount in component Ml(3). Neither light component contained ribose. 4. Alkaline phosphatase from Escherichia coli converted component Ml(2) into Ml(3). Incubation with crude preparations of phosphorylase b kinase or protein kinase in the presence of ATP converted component Ml(3) into Ml(2). 5. Phosphorylation of component Ml(3) with the kinases isolated from skeletal muscle and [gamma-(32)P]ATP gave incorporation of (32)P only into component Ml(2) whether whole myosin or separated low-molecular-weight components were used. 6. High-voltage electrophoresis at pH6.5 and pH1.8 of a chymotryptic digest of (32)P-labelled component Ml(2) yielded one major radioactive peptide containing serine phosphate. 7. The amino acid sequence of this peptide was shown to be: Arg-Ala-Ala-Ala-Glu-Gly-Gly-(Ser,Ser(P))-Asn-Val-Phe. This sequence shows no obvious similarity to the site phosphorylated in the conversion of phosphorylase b into phosphorylase a by phosphorylase b kinase. 8. Evidence suggests that in vivo all the 18500-molecular-weight light chain is in the phosphorylated form. The extent of dephosphorylation that occurred during myosin extraction depended on the conditions employed.
D000469:Alkaline Phosphatase; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D008722:Methods; D008970:Molecular Weight; D009132:Muscles / Q000032:analysis*; D009218:Myosins / Q000032:analysis*; D009578:Nitrobenzenes; D010455:Peptides / Q000032:analysis; D010758:Phosphorus / Q000032:analysis; D011817:Rabbits
D000596:Amino Acids; D009578:Nitrobenzenes; D010455:Peptides; D010758:Phosphorus; D000469:Alkaline Phosphatase; D009218:Myosins