A new hypothesis on the simultaneous direct and indirect proton pump mechanisms in NADH-quinone oxidoreductase (complex I)

Abstract 

Recently, Sazanov’s group reported the X-ray structure of whole complex I [Nature, 465, 441 (2010)], which presented a strong clue for a “piston-like” structure as a key element in an “indirect” proton pump. We have studied the NuoL subunit which has a high sequence similarity to Na⁺/H⁺ antiporters, as do the NuoM and N subunits. We constructed 27 site-directed NuoL mutants. Our data suggest that the H⁺/e⁻ stoichiometry seems to have decreased from (4H⁺/2e⁻) in the wild-type to approximately (3H⁺/2e⁻) in NuoL mutants. We propose a revised hypothesis that each of the “direct” and the “indirect” proton pumps transports 2H⁺ per 2e⁻.

Abbreviations: BN-PAGE, blue native gel electrophoresis, dNADH, deamino NADH, EIPA, 5-(N-ethyl-N-isopropyl) amiloride, Q, quinone, SQ, semiquinone, SQ_Nf, fast-relaxing semiquinone species, SQ_Ns, slow relaxing semiquinone species

Keywords: NADH-quinone oxidoreductase, Complex I, SQ_Nf-gated “direct” proton pump, Q_Ns-induced “indirect” proton pump, Conformationally-driven H⁺ pump