The mechanisms of reduction of protein mixed disulfides (dethiolation) in cardiac tissue.
Dethiolation of proteins (reduction of protein mixed disulfides) by NADPH-dependent and glutathione (GSH)-dependent enzymes, and by nonenzymatic reaction with GSH, was studied by electrofocusing methodology with glycogen phosphorylase b and creatine kinase as substrates. Phosphorylase b was not rapidly dethiolated by reduced glutathione alone, but a cardiac extract catalyzed rapid dethiolation by both an NADPH-dependent and a GSH-dependent process. In contrast, creatine kinase was actively dethiolated by GSH. This GSH-dependent dethiolation was not enhanced by a soluble extract of bovine heart. Creatine kinase was also not dethiolated by an NADPH-dependent process. Partial purification of the phosphorylase dethiolases showed that the NADPH-dependent dethiolase had both a high-molecular-weight and a low-molecular-weight component The properties of these components were similar to those of thioredoxin and thioredoxin reductase. These two components were sensitive to inhibition by phenylarsine oxide and inhibition was reversed by addition of a dithiol. In contrast, GSH-dependent dethiolation required a single component of low molecular weight. This process was less sensitive to phenylarsine oxide inhibition. These studies show that two cytosolic proteins, phosphorylase b and creatine kinase, were dethiolated by different mechanisms. Phosphorylase b was dethiolated by both NADPH-dependent and GSH-dependent enzymes found in a soluble extract of bovine heart. In contrast, creatine kinase was rapidly dethiolated nonenzymatically by GSH alone.
Park
E M
EM
Department of Biochemistry and Biophysics, Iowa State University, Ames 50011.
Thomas
J A
JA
D000818:Animals; D002417:Cattle; D003402:Creatine Kinase / Q000378:metabolism*; D004220:Disulfides / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism; D007700:Kinetics; D009132:Muscles / Q000201:enzymology; D009206:Myocardium / Q000201:enzymology*; D009249:NADP / Q000378:metabolism; D010084:Oxidation-Reduction; D010763:Phosphorylase b / Q000378:metabolism*; D006005:Phosphorylases / Q000378:metabolism*
D004220:Disulfides; D009249:NADP; D010763:Phosphorylase b; D006005:Phosphorylases; D003402:Creatine Kinase; D005978:Glutathione