Studies on the specificity of phosphorylase kinase using peptide substrates.
The action of phosphorylase kinase on synthetic peptides is reported. These peptides are variants of the amino acid sequence. Ser-Asp-Gln-Glu-Lys-Arg-Lys-Gln-Ile-Ser-Val-Arg-Gly-Leu, found in the natural substrate, phosphorylase b. The effects of size, the cluster of basic groups at the NH2-terminal side, the phosphorylatable seryl residue, the hydrophobic groups surrounding serine, and the arginyl function at the COOH-terminal side were tested and analyzed by evaluation of the kinetic parameters, Km and Vmax. The first 6 residues were found to be nonessential, but substitution of residues in the sequence. Lys-Gln-Ile-Ser-Val-Arg, had a large effect on phosphorylation. A comparison was made between the action of nonactivated and activated phosphorylase kinase on selected peptides and phosphorylase b. Various forms of phosphorylase b were tested as substrates for cyclic AMP-dependent protein kinase in the presence of effectors and salts. Although phosphorylase would not serve as a substrate for protein kinase, the aforementioned synthetic peptide of the phosphorylase b sequence would do so, indicating that the primary sequence surrounding the phosphorylatable serine did not block phosphorylation, which suggests that higher order structural features prohibit the phosphorylation.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D007700:Kinetics; D009132:Muscles / Q000201:enzymology; D010455:Peptides / Q000138:chemical synthesis*; D010764:Phosphorylase Kinase / Q000378:metabolism*; D011817:Rabbits; D013329:Structure-Activity Relationship
D000596:Amino Acids; D010455:Peptides; D010764:Phosphorylase Kinase