Affiliation:
1. the Department of Medicine, Division of Cardiology, The Johns Hopkins Medical Institutions, Baltimore, Md.
Abstract
Oxygen-derived free radicals have been reported to damage the sarcoplasmic reticulum (SR) Ca
2+
-ATPase, potentially contributing to cellular Ca
2+
overload and myocardial damage after ischemia and reperfusion. To determine whether the ATP binding site on Ca
2+
-ATPase is involved in oxygen radical injury, SR vesicles containing bound Ca
2+
-ATPase were isolated from rabbit cardiac and skeletal muscle and exposed to a hydroxyl radical (·OH)–generating system consisting of H
2
O
2
and Fe
3+
-nitrilotriacetic acid in amounts that generate a magnitude of ·OH similar to that which occurs in the reperfused heart. ·OH exposure completely inhibited Ca
2+
-ATPase activity and SR
45
Ca uptake for both cardiac and skeletal muscle. In contrast, when the purified vesicles were premixed with l mmol/L ATP before exposure to ·OH, complete protection was observed: there was no loss of ATPase activity or
45
Ca transport. No significant protection occurred with adenosine, sucrose, AMP, or ADP (l mmol/L each). SDS–gel electrophoresis indicated that ·OH did not damage the primary structure of the enzyme. Electron paramagnetic resonance spin-trapping experiments demonstrated that ATP did not scavenge ·OH. These results suggest that ·OH denatures the SR Ca
2+
-ATPase by directly attacking the ATP binding site, and occupation of the active site by ATP protects against ·OH-induced loss of enzymatic activity and SR Ca
2+
transport. The depletion of ATP that occurs during ischemia may enhance the toxic effect of ·OH at the time of reperfusion.
Publisher
Ovid Technologies (Wolters Kluwer Health)
Subject
Cardiology and Cardiovascular Medicine,Physiology
Cited by
238 articles.
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