Interaction of Peroxynitrite with Mitochondrial Cytochrome Oxidase

Abstract

Purified mitochondrial cytochrome c oxidase catalyzes the conversion of peroxynitrite to nitric oxide (NO). This reaction is cyanide-sensitive, indicating that the binuclear heme a 3 /Cu B center is the catalytic site. NO production causes a reversible inhibition of turnover, characterized by formation of the cytochrome a 3 nitrosyl complex. In addition, peroxynitrite causes irreversible inhibition of cytochrome oxidase, characterized by a decreased V max and a raised K m for oxygen. Under these conditions, the redox state of cytochrome a is elevated, indicating inhibition of electron transfer and/or oxygen reduction reactions subsequent to this center. The lipid bilayer is no barrier to these peroxynitrite effects, as NO production and irreversible enzyme inhibition were also observed in cytochrome oxidase proteoliposomes. Addition of 50 M peroxynitrite to 10 M fully oxidized enzyme induced spectral changes characteristic of the formation of ferryl cytochrome a 3 , partial reduction of cytochrome a, and irreversible damage to the Cu A site. Higher concentrations of peroxynitrite (250 M) cause heme degradation. In the fully reduced enzyme, peroxynitrite causes a red shift in the optical spectrum of both cytochromes a and a 3 , resulting in a symmetrical peak in the visible region. Therefore, peroxynitrite can both modify and degrade the metal centers of cytochrome oxidase.

The Peroxynitrite Reductase Activity of Cytochrome cOxidase Involves a Two-electron Redox Reaction at the Hemea 3-CuB Site

“…Ϫ with Oxidized Cytochrome Oxidase-Our current data concerning the consequences of the reaction between excess ONO 2 Ϫ and air-stable preparations of cytochrome c oxidase seem to be generally in accord with the findings of Sharpe and Cooper (20). In both studies the addition of excess oxidant has been found to lead to distinct spectral changes and irreversible inhibition of electron-transfer activity.…”

Endogenous Superoxide Production and the Nitrite/Nitrate Ratio Control the Concentration of Bioavailable Free Nitric Oxide in Leaves

“…Ascorbate is able to convert peroxynitrite into NO, thus regenerating free NO from the product of its reaction with superoxide (55,56). This reaction is further catalyzed by the presence of the mitochondrial enzyme, cytochrome c oxidase (56). We therefore suggest that an additional function of the high ascorbate concentrations in plants may be to preserve NO signaling pathways from variations in superoxide flux.…”

“…Interestingly we have recently described an alternative means of maintaining functional NO levels in the presence of superoxide. Ascorbate is able to convert peroxynitrite into NO, thus regenerating free NO from the product of its reaction with superoxide (55,56). This reaction is further catalyzed by the presence of the mitochondrial enzyme, cytochrome c oxidase (56).…”

Neuronal-glial Interactions Define the Role of Nitric Oxide in Neural Functional Processes

“…A NO-dependent reversible inhibition of cytochrome c oxidase modulating mitochondrial respiration has been reported in several cellular and subcellular systems, including isolated mitochondria, synaptosomes, macrophages, neurons and astrocytes [38-42]. Inhibition is due to NO competition with oxygen, but prolonged mitochondrial exposure to NO results in persistent inhibition of cytochrome c oxydase through formation of peroxynitrite [43-44]. Neurons are highly vulnerable to this type of mitochondrial inhibition, while astrocytes are much more resistant and they respond by activating mechanisms improving their survival and, as a consequence, also neuronal survival.…”