Spectral and redox characterization of the heme ci of the cytochrome b6f complex.

Absorption spectra of the purified cytochrome b(6)f complex from Chlamydomonas reinhardtii were monitored as a function of the redox potential. Four spectral and redox components were identified: in addition to heme f and the two b hemes, the fourth component must be the new heme c(i) (also denoted x) recently discovered in the crystallographic structures. This heme is covalently attached to the protein, but has no amino acid axial ligand. It is located in the plastoquinone-reducing site Q(i) in the immediate vicinity of a b heme. Each heme titrated as a one-electron Nernst curve, with midpoint potentials at pH 7.0 of -130 mV and -35 mV (hemes b), +100 mV (heme c(i)), and +355 mV (heme f). The reduced minus oxidized spectrum of heme c(i) consists of a broad absorption increase centered approximately 425 nm. Its potential has a dependence of -60 mV/pH unit, implying that the reduced form binds one proton in the pH 6-9 range. The Q(i) site inhibitor 2-n-nonyl-4-hydroxyquinoline N-oxide, a semiquinone analogue, induces a shift of this potential by about -225 mV. The spectrum of c(i) matches the absorption changes previously observed in vivo for an unknown redox center denoted "G." The data are discussed with respect to the effect of the membrane potential on the electron transfer equilibrium between G and heme b(H) found in earlier experiments.