Redox-Dependent Change of Nucleotide Affinity to the Active Site of the Mammalian Complex I

Abstract

A very potent and specific inhibitor of mitochondrial NADH:ubiquinone oxidoreductase (complex I), a derivative of NADH (NADH-OH) has recently been discovered (Kotlyar, A. B., Karliner, J. S., and Cecchini, G. (2005) FEBS Lett. 579, 4861-4866). Here we present a quantitative analysis of the interaction of NADH-OH and other nucleotides with oxidized and reduced complex I in tightly coupled submitochondrial particles. Both the rate of the NADH-OH binding and its affinity to complex I are strongly decreased in the presence of succinate. The effect of succinate is completely reversed by rotenone, antimycin A, and uncoupler. The relative affinity of ADPribose, a competitive inhibitor of NADH oxidation, is also shown to be significantly affected by enzyme reduction (K D of 30 and 500 μM for oxidized and the succinate-reduced enzyme, respectively). Binding of NADH-OH is shown to abolish the succinate-supported superoxide generation by complex I. Gradual inhibition of the rotenone-sensitive uncoupled NADH oxidase and the reverse electron transfer activities by NADH-OH yield the same final titration point (~0.1 nmol/mg of protein). The titration of NADH oxidase appears as a straight line, whereas the titration of the reverse reaction appears as a convex curve. Possible models to explain the different titration patterns for the forward and reverse reactions are briefly discussed.The mitochondrial proton-translocating NADH:ubiquinone oxidoreductase (complex I) and its prokaryotic homologues (NDH-1) 1 catalyze oxidation of NADH coupled to reduction of the membrane-located quinone. In aerobic organisms the enzymes provide at least two vitally important functions. First, they reoxidize NADH produced by multiple dehydrogenases, thus maintaining and controlling general oxidative metabolism. Second,

Mammalian Mitochondrial Complex I: Biogenesis, Regulation, and Reactive Oxygen Species Generation

“…This suggests that CI redox reactions can induce conformational changes. In its turn, nucleotide affinity to the CI active site appears to be redox sensitive (107).…”

Warm-preconditioning protects against acute heat-induced respiratory dysfunction and delays bleaching in a symbiotic sea anemone

“…As the TCA cycle is the primary source of NADH for mETC function (Berg et al, 2002) and assuming no increase in NAD + reduction by compensatory mechanisms, the decline in TCA cycle activity suggests potential NADH limitation and a reliance on SDH-generated FADH 2 as the source of electrons for the mETC. In other organisms, succinatedependent respiration can promote ROS generation through altered SDH activity (Jardim-Messeder et al, 2015), the autoxidation of partially reduced CoQ (Abele et al, 2007), reverse electron flow through NQO (Turrens and Boveris, 1980;Grivennikova et al, 2007) and -as SDH does not pump protons (Fig. 1) -changes to the IMM polarisation state.…”

Temporal changes in mitochondrial activities of rat heart after a single injection of iron, including increased complex II activity

“…Complex I can undergo conformational change depending on the relative amount of NAD ' versus NADH to facilitate reverse electron transfer from complex II (Grivennikova et al 2007). Thus, complex II links the TCA cycle to complex III of the electron transport chain and simultaneously restores reducing power to complex I, which was lost during the elimination of ROS caused by excessive iron.…”