Preferential binding of equine ferricytochrome c to the bacterial photosynthetic reaction center from Rhodobacter sphaeroides.
Redox titration of horse heart cytochrome c (cyt c), in the presence of varying concentrations of detergent-solubilized photosynthetic reaction center (RC) from Rhodobacter sphaeroides, revealed an RC concentration-dependent decrease in the measured cyt c midpoint potential that is indicative of a 3.6 +/- 0.2-fold stronger binding affinity of oxidized cytochrome to a single binding site. This effect was correlated with preferential binding in the functional complex by redox titration of the fraction of RCs exhibiting microsecond, first-order, special pair reduction by cytochrome. A binding affinity ratio of 3.1 +/- 0.4 was determined by this second technique, confirming the result. Redox titration of flash-induced intracomplex electron transfer also showed the association in the electron transfer-active complex to be strong, with a dissociation constant of 0.17 +/- 0.03 microM. The tight binding is associated with a slow off-rate which, in the case of the oxidized form, can influence the kinetics of P(+) reduction. The pitfalls of the common use of xenon flashlamps to photoexcite fast electron-transfer reactions are discussed with relation to the first electron transfer from primary to secondary RC quinone acceptors. The results shed some light on the diversity of kinetic behavior reported for the cytochrome to RC electron-transfer reaction.
Larson
J W
JW
Department of Biochemistry and Center for Biophysics and Computational Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.
Wraight
C A
CA
D000818:Animals; D003574:Cytochrome c Group / Q000378:metabolism*; D003902:Detergents; D004579:Electron Transport; D014450:Electron Transport Complex III / Q000378:metabolism; D005290:Ferric Compounds / Q000378:metabolism*; D006736:Horses; D008956:Models, Chemical; D010084:Oxidation-Reduction; D045322:Photosynthetic Reaction Center Complex Proteins / Q000378:metabolism*; D011199:Potentiometry; D011485:Protein Binding; D012242:Rhodobacter sphaeroides; D012995:Solubility; D017186:Titrimetry
D003574:Cytochrome c Group; D003902:Detergents; D005290:Ferric Compounds; D045322:Photosynthetic Reaction Center Complex Proteins; D014450:Electron Transport Complex III