Studies of the cytochrome bc1 complex and its electon acceptors in purple photosynthetic bacteria

The cytochrome bc1 complex is a key component of electron transfer chains in mitochondria and in many aerobic and photosynthetic bacteria. The bc1 complexes from photosynthetic bacteria provide many advantages for understanding electron transfer through the complex, as they are structurally simpler than the mitochondrial complexes and fast electron transfer after photoactivation of the reaction center can be easily followed using absorbance changes, something that cannot be done with mitochondria.

This dissertation focuses on three aspects related to the bc1 complexes: (1) The role of HiPIP and cytochrome cg, which are reduced by the bc1 complex, as alternative electron donors to the reaction center of Chromatium vinosum. Laser flash kinetics with intact Cm. vinosum cells showed that either HiPIP or cytochrome cg can be an efficient electron donor to the reaction center, depending on the media used to grow the cells. However, the preference for one electron donor over another does not arise from significant differences in protein abundance in the cells grown in the two different media. The mechanism of this "switch" remains to be elucidated. (2) Spectroscopic and oxidation-reduction properties of M183K and M183H variants of Rhodobacter capsulatus cytochrome c8. MCD and EPR spectra suggest that Rb. capsulatus cyt c\ is flexible and that one of the three histidines present outside the normal heme-binding domain can be recruited as an alternative to the methionine heme ligand found in the wild-type cytochrome. Titrations carried out in the oxidative direction differ markedly from those carried out in the reductive direction, mdicating the possible occurrence of redox-triggered conformational changes. (3) Interaction between Rb. capsulatus cytochrome bc1 and equine cytochrome c (a homolog for Rb. capsulatus cytochrome C2). Steady-state kinetic data, using site-specific cyt c1 mutants, showed that one acidic patch on the surface of cytochrome ci is unlikely to be involved in binding to cytochrome c. However, both steady-state kinetics and redox titration revealed that the phenylanaline at position 138 plays a critical role m maintaining a normal heme environment and suggest that this aromatic amino acid may participate in mediating electron transfer to and/or from the heme of cyt c1.