The overall objective of this research is to elucidate the pathway of electron transfer and the mechanism of energy transducing proton translocation in the cytochrome bc1 complex of the mitochondrial respiratory chain. During the forth-coming project period experimentation will be focused on identifying the proton translocation reactions and the redox proteins involved in those reactions.
The specific aims of the experimentation are to locate a ubiquinone and/or ubiquinol binding site, presumably on the iron-sulfur protein of the bc1 complex, from which protons are released coincident with electron transfer. Related studies will be conducted to test for a similar quinone/quinol binding site on cytochrome b.
A second aim of the experimentation will be to test whether cytochrome b is a transmembranous, redox linked proton pump. These studies will be carried out with mitochondria and with two species of aerobic bacteria which offer unique advantages to answering aspects of these questions which are not as readily answered by experimentation with mitochondria. The methods to be used include purification of membranous respiratory chain complexes into phospholipid vesicles, and photoaffinity labeling.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
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Physical Biochemistry Study Section (PB)
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Dartmouth College
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Hughes, Louise M; Lanteri, Charlotte A; O'Neil, Michael T et al. (2011) Design of anti-parasitic and anti-fungal hydroxy-naphthoquinones that are less susceptible to drug resistance. Mol Biochem Parasitol 177:12-9
Hughes, Louise M; Covian, Raul; Gribble, Gordon W et al. (2010) Probing binding determinants in center P of the cytochrome bc(1) complex using novel hydroxy-naphthoquinones. Biochim Biophys Acta 1797:38-43
Castellani, Michela; Covian, Raul; Kleinschroth, Thomas et al. (2010) Direct demonstration of half-of-the-sites reactivity in the dimeric cytochrome bc1 complex: enzyme with one inactive monomer is fully active but unable to activate the second ubiquinol oxidation site in response to ligand binding at the ubiquinone reducti J Biol Chem 285:502-10
Rottenberg, Hagai; Covian, Raul; Trumpower, Bernard L (2009) Membrane potential greatly enhances superoxide generation by the cytochrome bc1 complex reconstituted into phospholipid vesicles. J Biol Chem 284:19203-10
Ding, Martina G; di Rago, Jean-Paul; Trumpower, Bernard L (2009) Combining Inhibitor Resistance-conferring Mutations in Cytochrome b Creates Conditional Synthetic Lethality in Saccharomyces cerevisiae. J Biol Chem 284:8478-85
Zara, Vincenzo; Conte, Laura; Trumpower, Bernard L (2009) Evidence that the assembly of the yeast cytochrome bc1 complex involves the formation of a large core structure in the inner mitochondrial membrane. FEBS J 276:1900-14
Covian, Raul; Trumpower, Bernard L (2009) Ilicicolin Inhibition and Binding at Center N of the Dimeric Cytochrome bc1 Complex Reveal Electron Transfer and Regulatory Interactions between Monomers. J Biol Chem 284:8614-20
Covian, Raul; Trumpower, Bernard L (2009) The rate-limiting step in the cytochrome bc1 complex (Ubiquinol-Cytochrome c Oxidoreductase) is not changed by inhibition of cytochrome b-dependent deprotonation: implications for the mechanism of ubiquinol oxidation at center P of the bc1 complex. J Biol Chem 284:14359-67
Covian, Raul; Trumpower, Bernard L (2008) The dimeric structure of the cytochrome bc(1) complex prevents center P inhibition by reverse reactions at center N. Biochim Biophys Acta 1777:1044-52
Rotsaert, Frederik A J; Ding, Martina G; Trumpower, Bernard L (2008) Differential efficacy of inhibition of mitochondrial and bacterial cytochrome bc1 complexes by center N inhibitors antimycin, ilicicolin H and funiculosin. Biochim Biophys Acta 1777:211-9

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