Spectroscopic Studies of Mitochondrial Iron Sulfur Proteins (Succinate Dehydrogenase, Electron Paramagnetic Resonance, Fumarate Reductase).
Date of Award
Doctor of Philosophy (PhD)
Aerobic respiration is linked to oxidative phosphorylation and involves a redox-linked proton translocation across the mitochondrial inner membrane. Deeper understanding of the mechanism of mitochondrial electron transport requires detailed characterization of the redox centers present in respiratory enzyme complexes. A combination of low temperature MCD spectra and magnetization data, with parallel EPR, CD, and UV-visible experiments, has been used to characterize the structural, electronic, and magnetic properties of the iron sulfur centers in respiratory enzymes. The enzymes studied were those associated with electron transport in mammalian mitochondria, with comparative studies of similar enzymes and proteins in heterotropic bacteria. The results from studies of soluble high molecular weight NADH dehydrogenase are consistent with the presence of one 2Fe-2S and three 4Fe-4S centers, each approximately stoichiometric with FMN. Comparison of EPT spectra of Complex I and soluble enzyme indicates that the same clusters are present in the more intact particulate preparation. Spectroscopic experiments demonstrate that both yeast and mammalian succinate dehydrogenase, and the analogous fumarate reductase from E. coli, contain a 2Fe-2S , a 3Fe-xS , and a 4Fe-4S cluster in amounts stoichiometric with FAD. The trinuclear cluster was shown to be a necessary requirement for reconstitutive activity. In addition, evidence is presented to show that the 3Fe-xS and 4Fe-4S centers are distinct entities rather than interconversion products of the same cluster. Spectroscopic studies of the iron sulfur center in mammalian electron transfer flavoprotein (ETF) dehydrogenase were consistent with the presence of one 3Fe-4S cluster. In addition, dithionite reduction of ETF dehydrogenase was shown to involve addition of two electrons to FAD and one electron to the iron sulfur cluster, whereas the enzymatic process involves one electron reduction of both redox centers. Comparison of spectra from T. thermophilus Rieske protein with that from spinach ferredoxin indicated substantial differences in the electronic structure of their 2Fe-2S clusters. The differences most likely relate to variations in the ligand amino acid groups.
Morningstar, Joyce Elaine, "Spectroscopic Studies of Mitochondrial Iron Sulfur Proteins (Succinate Dehydrogenase, Electron Paramagnetic Resonance, Fumarate Reductase)." (1986). LSU Historical Dissertations and Theses. 4252.