title: Roles of semiquinone species in proton pumping mechanism by complex I.
abstract: Complex I (NDH-1) translocates protons across the membrane using electron transfer energy. Two different coupling mechanisms are currently being discussed for complex I: direct (redox-driven) and indirect (conformation-driven). Semiquinone (SQ) intermediates are suggested to be key for the coupling mechanism. Recently, using progressive power saturation and simulation techniques, three distinct SQ species were resolved by EPR analysis of E. coli complex I reconstituted into proteoliposomes. The fast-relaxing SQ (SQ(Nf)) signals completely disappeared in the presence of the uncoupler gramicidin D or the potent E. coli complex I inhibitor squamotacin. The slow-relaxing SQ (SQ(Ns)) signals were insensitive to gramicidin D, but they were sensitive to squamotacin. The very slow-relaxing SQ (SQ(Nvs)) signals were insensitive to both gramicidin D and squamotacin. Interestingly, no SQ(Ns) signal was observed in the ΔNuoL mutant, which lacks transporter module subunits NuoL and NuoM. Furthermore, we sought out the effect of using menaquinone (which has a lower redox potential compared to that of ubiquinone) as an electron acceptor on the proton pumping stoichiometry by in vitro reconstitution experiments with ubiquinone-rich or menaquinone-rich double knock-out membrane vesicles, which contain neither complex I nor NDH-2 (non-proton translocating NADH dehydrogenase). No difference in the proton pumping stoichiometry between menaquinone and ubiquinone was observed in the ΔNuoL and D178N mutants, which are considered to lack the indirect proton pumping mechanism. However, the proton pumping stoichiometry with menaquinone decreased by half in the wild-type. The roles and relationships of SQ intermediates in the coupling mechanism of complex I are discussed.
authors: Johnson Research Foundation, Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, 422 Curie Boulevard, Philadelphia, PA, 19104, USA, eikoo@mail.med.upenn.edu.
authors: Eiko
authors: E
authors: Nakamaru-Ogiso
authors: Madhavan
authors: M
authors: Narayanan
authors: Joseph A
authors: JA
authors: Sakyiama
mesh_terms: D015107:4-Butyrolactone / Q000031:analogs & derivatives / Q000737:chemistry; D000900:Anti-Bacterial Agents / Q000737:chemistry; D004578:Electron Spin Resonance Spectroscopy; D042967:Electron Transport Complex I / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D042963:Electron Transport Complex II / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006096:Gramicidin / Q000737:chemistry; D009245:NADH Dehydrogenase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010084:Oxidation-Reduction / Q000187:drug effects
chemical_list: D000900:Anti-Bacterial Agents; D029968:Escherichia coli Proteins; C102771:squamotacin; D006096:Gramicidin; D042963:Electron Transport Complex II; D009245:NADH Dehydrogenase; C479743:NuoL protein, E coli; C476247:NuoM protein, E coli; D042967:Electron Transport Complex I; D015107:4-Butyrolactone