title: Evidence for three separate electron flow pathways through Complex I: an inhibitor study.
abstract: The mammalian mitochondrial electron transport chain catalyzes the oxidation of NADH at pH 8.0 and pH 6.5, and the oxidation of NADPH at pH 6.5. The pH-dependencies of the rate of steady-state oxidation of NADPH and NADH by Complex I as well as by its flavoprotein fraction have been extensively studied by the laboratory of Hatefi. One model to explain these pH-dependent oxidations was proposed by Bakker and Albracht (Biochim. Biophys. Acta 850 (1986) 413-422 and 423-428, modified by Van Belzen and Albracht (Biochim. Biophys Acta 974 (1989) 311-320), which predicts that Complex I is a heterodimer with promoter B, containing FMN and Fe-S clusters 1-4 in stiochiometric amounts, catalyzing NADH oxidation at pH 8, and Protomer A, containing FMN and Fe-S clusters 2, 4, catalyzing NAD(P)H oxidation at pH 6.5. A pH-dependent transfer of electrons from protomer A Fe-S clusters 2, 4 to protomer B Fe-S clusters 2, 4 is an obligate step in the oxidation of NAD(P)H at low pH. Strict interpretation of this model allows for only three types of inhibitor: one which inhibits all three oxidase activities (type 1); one which inhibits NADH oxidase, pH 8.0 (type 4) and a third which inhibits NAD(P)H oxidase, pH 6.5 (type 5). Another possibility is that there are three separate pathways of oxidation of NAD(P)H, which would allow for a total of seven different types of inhibitor, e.g., the three types above plus type 2 inhibiting NADH oxidase pH 8.0 and pH 6.5; type 3 inhibiting NADH oxidase pH 8.0, and NADPH oxidase pH 6.5; type 6 inhibiting NADH oxidase pH 6.5; and type 7 inhibiting NADPH oxidase pH 6.5. Using a series of thirteen inhibitors of Complex I activity and the chemical modification reagent ethoxyformic anhydride (EFA), four different inhibitor types were found: seven inhibitors of type 1, four inhibitors of type 2, one inhibitor of type 3 and one inhibitor of type 4. Treatment of submitochondrial particles (SMP) with EFA abolished NADH-dependent reduction of coenzyme Q at both pH 8.0 and 6.5, while inhibiting NADPH-dependent reduction of coenzyme Q at pH 6.5 by only 30%. These results do not support the heterodimer model of Complex I electron transport of Bakker and Albracht, but do support three separate electron flow pathways through complex 1 from reduced pyridine nucleotides to coenzyme Q. A new model of electron flow through Complex I based on these finding is proposed.
authors.affiliation: Indiana University School of Medicine, Northwest Center for Medical Education, Gary 46408, USA.
authors.forename: W M
authors.initials: WM
authors.lastname: Anderson
authors.forename: D
authors.initials: D
authors.lastname: Trgovcich-Zacok
mesh_terms: D000818:Animals; D002417:Cattle; D004047:Diethyl Pyrocarbonate / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004579:Electron Transport; D006863:Hydrogen-Ion Concentration; D008929:Mitochondria, Heart / Q000201:enzymology / Q000378:metabolism*; D008954:Models, Biological; D009243:NAD / Q000378:metabolism; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009249:NADP / Q000378:metabolism; D013367:Submitochondrial Particles / Q000201:enzymology / Q000378:metabolism*; D017186:Titrimetry
chemical_list: D009243:NAD; D009249:NADP; D016660:NAD(P)H Dehydrogenase (Quinone); D004047:Diethyl Pyrocarbonate