title: Active/de-active state transition of the mitochondrial complex I as revealed by specific sulfhydryl group labeling.
abstract: The sensitivities of NADH oxidase and/or NADH-ubiquinone reductase activities of submitochondrial particles and purified complex I towards N-ethylmaleimide (NEM) and other SH-reagents were studied. Only thermally de-activated preparations IA.D. Vinogradov (1998) Biochim. Biophys. Acta 1364, 169-185] were inhibited by SH-reagents whereas the redox-pulsed, activated enzyme was resistant to the inhibitors. The pH profile of the pseudo-first order inhibition rate suggested a pKa of about 10 for the de-activation-dependent, NEM-reactive sulfhydryl group. NADH-ubiquinone reductase of activated particles treated with an excess of NEM followed by removal of the inhibitor was still capable of slow reversible active/de-active transition. When active, NEM-treated particles were de-activated and further inhibited by N-fluorescein maleimide, specific incorporation of the fluorescence label into low molecular mass polypeptide was evident. Comparison of the specific fluorescence labeling of submitochondrial particles, crude and purified complex I showed that the active/de-active state-dependent SH-group is located in a 15 kDa polypeptide (most likely in the 15 kDa IP subunit of the iron-sulfur protein-containing fraction of complex I).
authors.affiliation: Department of Biochemistry, School of Biology, Moscow State University, Russia.
authors.forename: E V
authors.initials: EV
authors.lastname: Gavrikova
authors.forename: A D
authors.initials: AD
authors.lastname: Vinogradov
mesh_terms: D000818:Animals; D002417:Cattle; D004591:Electrophoresis, Polyacrylamide Gel; D008929:Mitochondria, Heart / Q000201:enzymology*; D015335:Molecular Probes; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013367:Submitochondrial Particles / Q000201:enzymology; D013438:Sulfhydryl Compounds / Q000378:metabolism*
chemical_list: D015335:Molecular Probes; D013438:Sulfhydryl Compounds; D016660:NAD(P)H Dehydrogenase (Quinone)