We propose to study the mechanism of electron and proton transfer pathways in mitochondrial NADH-quinone oxidoreductase (often referred to as Complex I). this complex plays a central role in the oxidation of NADH, the reducing product of cellular metabolism, by the respiratory chain. Complex I is the most complicated and least understood energy transducing proton-motive enzyme of the respiratory chain. the proposed research takes new directions in the investigation of the mechanism, kinetics, and regulatory properties of this complex in both the isolated and membrane-bound states. The specific studies will include: [I] determination of the minimal structure of Complex I capable of catalyzing NADH oxidation which is coupled with vectorial transfer of protons across the membrane; [II] determination of the internal electron transfer sequence within intrinsic redox components of NADH-quinone oxidoreductase by utilizing artificial electron acceptors (transition metal complexes) with proper redox potentials and appropriate other parameters (hydrophilicity, charge, size); [III] physico-chemical studies on the ubisemiquinones associated with the specific binding sites in Complex I which was recently discovered in Russian Co-P.I.'s laboratory; [IV] studies of molecular events involved in the slow active/inactive transition of Complex I, rediscovered recently by the Russian Co-P.I., which will be extended to searches of physiologically relevant factors which are involved in the control of hysteretic behavior of this complex. The P.I. has extensive experience with EPR studies and with thermodynamic analysis of the intrinsic paramagnetic redox centers of mitochondrial and microbial systems. Specifically, the P.I. has made significant contribution in the characterization of iron-sulfur clusters, flavin and ubiquinone free radicals. The Russian Co-P.I. is an established biochemist and has recently developed unique submitochondrial particle preparations having tightly coupled Site I energy transduction. The collaborative program between these investigators, therefore, is expected to greatly enhance the understanding of the mechanism of energy coupling and its regulation.

Agency
National Institute of Health (NIH)
Institute
Fogarty International Center (FIC)
Type
Small Research Grants (R03)
Project #
1R03TW000140-01A2
Application #
3432585
Study Section
Special Emphasis Panel (SRC)
Project Start
1993-07-01
Project End
1996-06-30
Budget Start
1993-07-01
Budget End
1994-06-30
Support Year
1
Fiscal Year
1993
Total Cost
Indirect Cost
Name
University of Pennsylvania
Department
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Magnitsky, Sergey; Toulokhonova, Larisa; Yano, Takahiro et al. (2002) EPR characterization of ubisemiquinones and iron-sulfur cluster N2, central components of the energy coupling in the NADH-ubiquinone oxidoreductase (complex I) in situ. J Bioenerg Biomembr 34:193-208
Gavrikova, E V; Vinogradov, A D (1999) Active/de-active state transition of the mitochondrial complex I as revealed by specific sulfhydryl group labeling. FEBS Lett 455:36-40
Vinogradov, A D; Gavrikova, E V; Grivennikova, V G et al. (1999) Catalytic properties of mitochondrial NADH-ubiquinone reductase (Complex I). Biochemistry (Mosc) 64:136-52
Galkin, A S; Grivennikova, V G; Vinogradov, A D (1999) -->H+/2e- stoichiometry in NADH-quinone reductase reactions catalyzed by bovine heart submitochondrial particles. FEBS Lett 451:157-61
Zakharova, N V; Zharova, T V; Vinogradov, A D (1999) Kinetics of transhydrogenase reaction catalyzed by the mitochondrial NADH-ubiquinone oxidoreductase (Complex I) imply more than one catalytic nucleotide-binding sites. FEBS Lett 444:211-6
Ushakova, A V; Grivennikova, V G; Ohnishi, T et al. (1999) Triton X-100 as a specific inhibitor of the mammalian NADH-ubiquinone oxidoreductase (Complex I). Biochim Biophys Acta 1409:143-53
Vinogradov, A D (1998) Catalytic properties of the mitochondrial NADH-ubiquinone oxidoreductase (complex I) and the pseudo-reversible active/inactive enzyme transition. Biochim Biophys Acta 1364:169-85
Grivennikova, V G; Maklashina, E O; Gavrikova, E V et al. (1997) Interaction of the mitochondrial NADH-ubiquinone reductase with rotenone as related to the enzyme active/inactive transition. Biochim Biophys Acta 1319:223-32
Zharova, T V; Vinogradov, A D (1997) A competitive inhibition of the mitochondrial NADH-ubiquinone oxidoreductase (complex I) by ADP-ribose. Biochim Biophys Acta 1320:256-64
Gavrikova, E V; Grivennikova, V G; Sled, V D et al. (1995) Kinetics of the mitochondrial three-subunit NADH dehydrogenase interaction with hexammineruthenium(III). Biochim Biophys Acta 1230:23-30

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