Comparison of electrostatic interactions and of protein-protein orientations in electron-transfer reactions of plastocyanin with the triplet state of zinc cytochrome c and with zinc cytochrome c cation radical.

Abstract

Photoinduced reduction of cupriplastocyanin by the triplet state of zinc cytochrome c (the "forward" reaction) and the subsequent thermal oxidation of cuproplastocyanin by zinc cytochrome c cation radical (the "back" reaction) at ionic strengths from 40 mM to 3.00 M are studied by laser kinetic spectroscopy (so-called flash photolysis). Variation of the bimolecular rate constants over the entire range of ionic strength cannot be explained in terms of monopole-monopole interactions between the protein molecules, but it can be explained in terms of monopole-monopole, monopole-dipole, and dipole-dipole interactions. Analysis of the kinetic results in terms of these electrostatic interactions reveals the overall protein-protein orientation for electron transfer. In both the forward and back reactions the exposed heme edge in zinc cytochrome c apparently abuts the negatively-charged (acidic) patch on the plastocyanin surface, which is remote from the copper atom, and not the electroneutral (hydrophobic) patch, which is proximate to the copper atom. The acidic patch is large, and this analysis cannot rule out a relatively small difference in protein-protein orientations for the forward and back reactions. These two reactions are compared with the previously studied reduction of cupriplastocyanin by ferrocytochrome c. Although native cytochrome c and its zinc derivative have very similar structural and electrostatic properties, the reactive forms of the cytochrome c/plastocyanin and zinc cytochrome c/plastocyanin complexes may adopt somewhat different protein-protein orientations or may adopt similar orientations but differ in dynamic properties.