Interactions between nitric oxide and peroxynitrite during prostaglandin endoperoxide H synthase-1 catalysis: A free radical mechanism of inactivation

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• 作者：André ; s Trostchansky ; Valerie B. O'Donnell ; Douglas C. Goodwin ; Lisa M. L ; ino ; Lawrence J. Marnett ; Rafael Radi ; Homero Rubbo
• 关键词：PGHS-1 ; Arachidonic acid ; Peroxynitrite ; Nitric oxide ; Tyrosine nitration ; Peroxidase ; Cyclooxygenase
• 刊名：Free Radical Biology and Medicine
• 出版年：2007
• 期刊代码：105_08915849
• 类别：med
• 出版时间：1 April 2007
• 卷：42
• 期：7
• 页码：1029-1038
• 文件大小：504 K

摘要

Peroxynitrite (ONOO^–) can serve either as a peroxide substrate or as an inactivator of prostaglandin endoperoxide H synthase-1 (PGHS-1). Herein, the mechanism of PGHS-1 inactivation by ONOO^– and the modulatory role that nitric oxide (NO) plays in this process were studied. PGHS-1 reacted with ONOO^– with a second-order rate constant of 1.7 × 10⁷ M^{− 1} s^{− 1} at pH 7.0 and 8 °C. In the absence of substrates, the enzyme was dose-dependently inactivated by ONOO^– in parallel with 3-nitrotyrosine formation. However, when PGHS-1 was incubated with ONOO^– in the presence of substrates, the direct reaction with ONOO^– was less relevant and ONOO^–-derived radicals became involved in enzyme inactivation. Bicarbonate at physiologically relevant concentrations enhanced PGHS-1 inactivation and nitration by ONOO^–, further supporting a free radical mechanism. Importantly, NO (0.4–1.5 μM min^{− 1}) was able to spare the peroxidase activity of PGHS-1 but it enhanced ONOO^–-mediated inactivation of cyclooxygenase. The observed differential effects of NO on ONOO^–-mediated PGHS-1 inactivation emphasize a novel aspect of the complex modulatory role that NO plays during inflammatory processes. We conclude that ONOO^–-derived radicals inactivate both peroxidase and cyclooxygenase activities of PGHS-1 during enzyme turnover. Finally, our results reconcile the proposed alternative effects of ONOO^– on PGHS-1 (activation versus inactivation).