Hydroxyl radical inhibits sarcoplasmic reticulum Ca2+-ATPase function by direct attack on the ATP binding site

Kai Y. Xu, Jay L. Zweier, Lewis C. Becker

Research output: Contribution to journalArticlepeer-review

Abstract

Oxygen-derived free radicals have been reported to damage the sarcoplasmic reticulum (SR) Ca2+-ATPase, potential contributing to cellular Ca2+ overload and myocardial damage after ischemia and reperfusion. To determine whether the ATP binding site on Ca2+-ATPase is involved in oxygen radical injury, SR vesicles containing bound Ca2+-ATPase were isolated from rabbit cardiac and skeletal muscle and exposed to a hydroxyl radical ((·OH)- generating system consisting of H2O2 and Fe2+-nitriloacetic acid in amounts that generate a magnitude of ·OH similar to that which occurs in the reperfused heart. ·OH exposure completely inhibited Ca2+-ATPase activity and SR 45Ca uptake for both cardiac and skeletal muscle. In contrast, when the purified vesicles were premixed with 1 mmol/I. ATP before exposure to ·OH, complete protection was observed; there was no loss of ATPase activity or 45Ca transport. No significant protection occurred with adenosine, sucrose, AMP, or ADP (1 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 Ca2+- 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 Ca2+ transport. The depletion of ATP that occurs during ischemia may enhance the toxic effect of ·OH at the time of reperfusion.

Original languageEnglish (US)
Pages (from-to)76-81
Number of pages6
JournalCirculation research
Volume80
Issue number1
DOIs
StatePublished - 1997

Keywords

  • ATP
  • Ca-ATPase
  • hydroxyl radical
  • ischemia/reperfusion
  • sarcoplasmic reticulum

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine

Fingerprint Dive into the research topics of 'Hydroxyl radical inhibits sarcoplasmic reticulum Ca<sup>2+</sup>-ATPase function by direct attack on the ATP binding site'. Together they form a unique fingerprint.

Cite this