Lack of Oxygen Deactivates Mitochondrial Complex I

Abstract

For S-nitrosothiols and peroxynitrite to interfere with the activity of mitochondrial complex I, prior transition of the enzyme from its active (A) to its deactive, dormant (D) state is necessary. We now demonstrate accumulation of the D-form of complex I in human epithelial kidney cells after prolonged hypoxia. Upon reoxygenation after hypoxia there was an initial delay in the return of the respiration rate to normal. This was due to the accumulation of the D-form and its slow, substratedependent reconversion to the A-form. Reconversion to the A-form could be prevented by prolonged incubation with endogenously generated NO. We propose that the hypoxic transition from the A-form to the D-form of complex I may be protective, because it would act to reduce the electron burst and the formation of free radicals during reoxygenation. However, this may become an early pathophysiological event when NO-dependent formation of S-nitrosothiols or peroxynitrite structurally modifies complex I in its D-form and impedes its return to the active state. These observations provide a mechanism to account for the severe cell injury that follows hypoxia and reoxygenation when accompanied by NO generation.The mechanisms underlying the cellular response to hypoxia and their consequences are not completely understood. Because the mitochondrial respiratory chain is the major consumer of oxygen, mitochondria are likely to play a significant role in regulating its distribution in cells and tissues (1). Cells have the ability to decrease oxygen demand at low [O 2 ] (2, 3), and the affinity of cytochrome c oxidase for oxygen is considered to be the most important factor in the decrease of mitochondrial oxygen consumption during hypoxia (4, 5). The interaction of nitric oxide (NO) 2 with cytochrome c oxidase has been shown to be a significant determinant of the affinity of this enzyme for oxygen and is responsible for reducing cellular consumption of oxygen at low [O 2 ]. Nitric oxide also plays a role in the early reduction of mitochondrial cytochromes that occurs as the [O 2 ] decreases (6, 7). A direct consequence of such reduction is a backlog of electrons at all the redox centers of the respiratory chain, including cytochromes and ubiquinone, as well as the intramitochondrial pool of NAD(P)H.Mitochondrial complex I (EC 1.6.5.3, proton-translocating NADH:ubiquinone oxidoreductase) is responsible for oxidation of matrix NADH by membrane-bound ubiquinone and is the major entry point for electrons to the respiratory chain (8). It is also a major source of mitochondrial reactive oxygen species (ROS) (9 -11). Two catalytically and structurally distinct forms of complex I have been identified in partially purified preparations in vitro: one is a fully competent, "active" A-form, and the other is a dormant, silent, "deactivated" D-form (12). In such systems a so-called pseudo-reversible A/D transition has been described in mammalian and other vertebrate complex I (13). The turnover number of active complex I in submitochondrial particles (SM...

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