The aims of this research which deals with non-cytochrome-containing components are to obtain basic information on the respiratory chain enzyme system of mitochondrial inner membrane so as to enable conclusions concerning obligatory carriers, their sequence, actions, mechanisms and subjects related to the electron transport chain, as well as to determine the role of the membrane in these processes, utilizing the methods of sequential resolution, systematic reconstitution, identification and characterization of the isolated enzymes. First, the resolved enzymes and components will be purified and then rigorously characterized by all conceivable physico-chemical and biochemical techniques, including reaction kinetics, amino acid sequence, EPR, ENDOR, crystallization, EXAFS and SANES, among others. Amino acid sequence determination is most definitive in delineating the identity or non-identity of the components. However, it must be emphasized that reconstitution will be used as the final and most reliable method to ascertain that no degradation or changes occurred during extensive purification of the components. At the same time, purposeful change or modification, based on well-reasoned design, of the isolated enzymes will be used to examine the reconstitutive activity in order to ascertain how much leeway nature allows for deviation from normal function. However, an ultimate goal is reconstitution from well defined components of the complete electron transfer apparatus with all functional manifestations of the mitochondrion. It should be pointed out that this application (which differs from another project) emphasizes and deals with aspects of the non-cytochrome-containing components of the respiratory chain.
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| Clarkson, G H; King, T E; Lindsay, J G (1987) Biosynthesis and processing of the large and small subunits of succinate dehydrogenase in cultured mammalian cells. Biochem J 244:15-20 |
| King, T E; Kim, C H (1986) Preparation of hinge protein and its requirement for interaction of cytochrome c with cytochrome c1. Methods Enzymol 126:238-53 |
