Electron-transfer reactions of photoreduced flavin analogs with c-type cytochromes: quantitation of steric and electrostatic factors

Abstract

Correlations were found between rate constants and the difference in redox potential of the reactants for electron-transfer reactions between oxidized cytochromes and either photoproduced riboflavin or FMN semiquinones (the latter rate constants extrapolated to infinite ionic strength). The riboflavin-cytochrome rate constants are about 70% of those for reduction by lumiflavin, probably because of steric interference by the ribityl side chain. Reduction of cytochromes by FMN semiquinone was ionic strength dependent in all cases, due to electrostatic interactions. Extrapolation of rate constants to infinite ionic strength shows that the phosphate exerts a significant steric effect as well (rate constants average .apprx. 27% of those for lumiflavin, although part of this decrease is due to a difference in the semiquinone pK value). Differences in the magnitude of the FMN steric effect correlate well with surface topology differences for those cytochromes whose 3-dimensional structures are known. Mitochondrial cytochromes c and the cytochromes c2 all showed attractive (plus-minus) interaction with FMN in spite of the fact that some of these proteins have large net negative charges. Four small c-type cytochromes (including Pseudomonas cytochrome c-551) show a weak repulsive interaction with FMN semiquinone. Flavosemiquinones probably interact at a site on the cytochromes that is near the exposed heme edge. There is a large positive electrostatic field at this site in mitochondrial cytochrome c and the cytochromes c2, but this region is primarily hydrophobic in Pseudomonas cytochrome c-551 and in the other small bacterial cytochromes. In the Pseudomonas reaction, FMN interacts weakly with a negative electrostatic field, which must be somewhat removed from the site of electron transfer. The relative contributions of redox potential, steric effects and electrostatic interactions for the flavosemiquinone-cytochrome reactions described herein are roughly equivalent and correspond to rate constant changes of 2- to 5-fold.