Defining the roles of protein-protein and protein-metal interactions in biological electron transfer processes.

Abstract

Biological electron transfer is a fundamental process for living systems. Aerobic respiration and photosynthesis are two energy transducing biological electron transfer pathways. A knowledge of the factors that govern specificity and rates of electron transfer is essential to understand this process. Two blue copper/c-type cytochrome electron transfer couples (azurin/cytochrome c551 from Pseudomonas aeruginosa and plastocyanin/cytochrome c552(3) from cyanobacteria and higher plants) were selected for study representing respiration and photosynthesis with the aim of obtaining information on protein-protein recognition and protein-metal interactions. Plastocyanin from Lycoperison esculentum (tomato) was cloned and sequenced in preparation for site directed mutagensis studies on the metal center and on surface residues important for docking with physiological partners. Zn(II) cytochrome c551 from Pseudomonas aeruginosa was prepared, characterized and used in binding constant studies with its proposed physiological partner, azurin. These studies showed the binding between Zn(II) c551 and Cu(II) azurin to be weak at $<$10$\sp4$M$\sp{-1}$. This weak binding may indicate that these proteins are not physiological partners but rather, interchangeable components. The photodecay process that Zn(II) c-type cytochromes have been known to undergo was studied. A decomposition scheme was proposed which involves molecular oxygen, proceeding through a porphyrin radical cation and ending in a covalently modified porphyrin. 2 dimensional NMR studies on Fe(II) c551 resulted in the spin system and sequential assignments for this protein as a first step towards comparing Zn(II) and Fe(II) c551 structures and the effect metal substitution.