The aim of this research is to elucidate the protein structural requirements for electron transfer and control in the mitochondrial electron transfer chain, and in the microsomal NADPH cytochrome c (P450) reductase system. It is proposed that control of oxygen consumption at the level of cytochrome oxidase may involve a mechanism similar to that defined for hemoglobin, in which changes in electrostatic interactions between subunits determine the affinity of oxygen binding at the heme. In the case of cytochrome oxidase, the strong ionic interactions with its substrate, cytochrome c, are likely to influence the conformation of the oxidase, as is the membrane potential gradient. The effects of the presence, absence and binding affinity of cytochrome c on respiratory control in coupled inner mitochondrial membranes will be studied. Steady-state, pre-steady state, and low-temperature kinetic analyses of the reactions of native and modified cytochromes c with cytochrome c oxidase will be carried out with a purified, mono disperse oxidase preparation, as well as with the membrane-bound form, in order to further clarify the mechanism and mechanics of the reaction. This will also require direct binding studies and an investigation of the kinetic effects of covalently cross-linking cytochrome c to purified and membrane-bound oxidase. The reaction of cytochrome c with microsomal NADPH-cytochrome c (P450) reductase will be investigated as a function of cytochrome c structure, ionic strength, presence of cytochrome P450, and the type of reductase preparation (protease-solubilized, detergent solubilized, and membrane-bound), in order to define the nature of the protein interactions involved.

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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Michigan State University
Schools of Arts and Sciences
East Lansing
United States
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Hiser, Carrie; Liu, Jian; Ferguson-Miller, Shelagh (2018) The K-path entrance in cytochrome c oxidase is defined by mutation of E101 and controlled by an adjacent ligand binding domain. Biochim Biophys Acta Bioenerg 1859:725-733
Liu, Jian; Hiser, Carrie; Ferguson-Miller, Shelagh (2017) Role of conformational change and K-path ligands in controlling cytochrome c oxidase activity. Biochem Soc Trans 45:1087-1095
Liu, Jian; Hiser, Carrie; Ferguson-Miller, Shelagh (2017) Correction: Role of conformational change and K-path ligands in controlling cytochrome c oxidase activity. Biochem Soc Trans 45:1345
Li, Fei; Liu, Jian; Liu, Nan et al. (2016) Translocator Protein 18 kDa (TSPO): An Old Protein with New Functions? Biochemistry 55:2821-31
Li, Fei; Liu, Jian; Zheng, Yi et al. (2015) Protein structure. Crystal structures of translocator protein (TSPO) and mutant mimic of a human polymorphism. Science 347:555-8
Li, Fei; Liu, Jian; Valls, Lance et al. (2015) Identification of a key cholesterol binding enhancement motif in translocator protein 18 kDa. Biochemistry 54:1441-3
Li, Fei; Liu, Jian; Garavito, R Michael et al. (2015) Evolving understanding of translocator protein 18 kDa (TSPO). Pharmacol Res 99:404-9
Li, Fei; Liu, Jian; Zheng, Yi et al. (2015) Response to Comment on ""Crystal structures of translocator protein (TSPO) and mutant mimic of a human polymorphism"". Science 350:519
Schwaighofer, Andreas; Ferguson-Miller, Shelagh; Naumann, Renate L C et al. (2014) Phase-sensitive detection in modulation excitation spectroscopy applied to potential induced electron transfer in cytochrome c oxidase. Appl Spectrosc 68:5-13
Buhrow, Leann; Hiser, Carrie; Van Voorst, Jeffrey R et al. (2013) Computational prediction and in vitro analysis of potential physiological ligands of the bile acid binding site in cytochrome c oxidase. Biochemistry 52:6995-7006

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