title: Effect of sulfhydryl-disulfide state on protein phosphorylation: phosphorylation of bovine serum albumin.
abstract: Bovine serum albumin (BSA) was phosphorylated by the catalytic subunit of cAMP-dependent protein kinase under general protein phosphorylation conditions. The optimal pH for this phosphorylation was 9.0. The K0.5 (the concentration required for 50% of maximal phosphorylation) for BSA at pH 7.5 was 15 microM. One mole of phosphate was incorporated per mole of BSA, and only one phosphopeptide fragment was obtained after extensive proteolysis with trypsin. BSA phosphorylation required dithiothreitol or GSH, but GSH was only one-fiftieth as effective as dithiothreitol. GSSG counteracted the effect of dithiothreitol and GSH. Phosphorylation increased in a time-dependent and dithiothreitol concentration-dependent manner when BSA was preincubated with dithiothreitol. The increase in the incorporation of 32P correlated with the appearance of up to six free sulfhydryl groups. The effect of dithiothreitol on BSA appeared reversible, since reoxidation of reduced BSA decreased its susceptibility to phosphorylation. These experiments showed that this in vitro phosphorylation is dependent on the sulfhydryl-disulfide state of BSA. The possible implications of the sulfhydryl-disulfide state of proteins in the regulation of phosphorylation are discussed.
authors.forename: S L
authors.initials: SL
authors.lastname: Chen
authors.forename: K H
authors.initials: KH
authors.lastname: Kim
mesh_terms: D000818:Animals; D002384:Catalysis; D002417:Cattle; D000242:Cyclic AMP / Q000378:metabolism; D004220:Disulfides / Q000378:metabolism; D004229:Dithiothreitol / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D010084:Oxidation-Reduction; D010766:Phosphorylation; D011494:Protein Kinases / Q000494:pharmacology*; D012710:Serum Albumin, Bovine / Q000378:metabolism*; D013438:Sulfhydryl Compounds / Q000378:metabolism*
chemical_list: D004220:Disulfides; D013438:Sulfhydryl Compounds; D012710:Serum Albumin, Bovine; D000242:Cyclic AMP; D011494:Protein Kinases; D004229:Dithiothreitol