Free radical inactivation of rabbit muscle creatinine kinase: catalysis by physiological and hydrolyzed ICRF-187 (ICRF-198) iron chelates.
Creatine kinase is a sulfhydryl containing enzyme that is particularly susceptible to oxidative inactivation. This enzyme is potentially vulnerable to inactivation under conditions when it would be used as a diagnostic marker of tissue damage such as during cardiac ischemia/reperfusion or other oxidative tissue injury. Oxidative stress in tissues can induce the release of iron from its storage proteins, making it an available catalyst for free radical reactions. Although creatinine kinase inactivation in a heart reperfusion model has been documented, the mechanism has not been fully described, particularly with regard to the role of iron. We have investigated the inactivation of rabbit muscle creatine kinase by hydrogen peroxide and by xanthine oxidase generated superoxide or Adriamycin radicals in the presence of iron catalysts. As shown previously, creatine kinase was inactivated by hydrogen peroxide. Ferrous iron enhanced the inactivation. In addition, micromolar levels of iron and iron chelates that were reduced and recycled by superoxide or Adriamycin radicals were effective catalysts of creatinine kinase inactivation. Of the physiological iron chelates studied, Fe(ATP) was an especially effective catalyst of inactivation by what appeared to be a site-localized reaction. Fe(ICRF-198), a non-physiological chelate of interest because of its putative role in alleviating Adriamycin-induced cardiotoxicity, also catalyzed the inactivation. Scavenger studies implicated hydroxyl radical as the oxidant involved in iron-dependent creatine kinase inactivation. Loss of protein thiols accompanied loss of creatine kinase activity. Reduced glutathione (GSH) provided marked protection from oxidative inactivation, suggesting that enzyme inactivation under physiological conditions would occur only after GSH depletion.
Thomas
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Department of Pathology, Christchurch School of Medicine, New Zealand.
Carr
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Winterbourn
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D000818:Animals; D002374:Catalase / Q000494:pharmacology; D002384:Catalysis; D003402:Creatine Kinase / Q000302:isolation & purification*; D004317:Doxorubicin / Q000494:pharmacology; D005029:Ethylenediamines / Q000494:pharmacology*; D005609:Free Radicals / Q000494:pharmacology; D005998:Glycine / Q000031:analogs & derivatives* / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000494:pharmacology; D006868:Hydrolysis; D007501:Iron / Q000494:pharmacology; D007502:Iron Chelating Agents / Q000494:pharmacology*; D007527:Isoenzymes; D007700:Kinetics; D009132:Muscles / Q000201:enzymology*; D018384:Oxidative Stress; D011817:Rabbits; D011929:Razoxane / Q000494:pharmacology*; D013481:Superoxides / Q000494:pharmacology; D014969:Xanthine Oxidase / Q000378:metabolism
D005029:Ethylenediamines; D005609:Free Radicals; D007502:Iron Chelating Agents; D007527:Isoenzymes; D013481:Superoxides; D011929:Razoxane; C059866:ICRF 198; D004317:Doxorubicin; D006861:Hydrogen Peroxide; D007501:Iron; D002374:Catalase; D014969:Xanthine Oxidase; D003402:Creatine Kinase; D005998:Glycine