PH-Dependent interconversion between the two redox forms of chloroplast cytochrome b-559

Abstract

Spinach chloroplast cytochrome b-559 is characterized (cf. Ref. 1) by exhibiting both a pH-independent hydroquinone-reducible high-potential form (midpoint potential U′ 0, pH 7 = 340 mV) and a pH-dependent (p K a = 7.6) dithionite-reducible low-potential form (midpoint potential U′ 0, pH 7 = 135 mV). Untreated fresh chloroplasts present about two-thirds of their cytochrome b-559 in its high-potential form, which is rapidly reduced by hydroquinone. However, fresh chloroplasts treated in the light with a high concentration of the protonophoric uncoupler carbonylcyanide- p-trifluoromethoxy-phenylhydrazone (FCCP) contain only the low-potential form not reducible by hydroquinone, a change that is brought about by the irreversible conversion of the high-potential form into the low-potential one. In the presence of a lower concentration of the uncoupler, the high-potential form likewise becomes transformed into the low-potential form, but remarkably this can then be subsequently reduced by hydroquinone, although only at a comparatively slow rate, thus indicating reversibility of the conversion process between both redox couples. These results, which are discussed in detail in a broad context, reinforce our view (cf. Ref. 1) that cytochrome b-559 operates not only as a redox but also as an acid-base energy-transducing system, of both the energized oxidized- and basic-form type at two alternate potentials and p K a 's. Accordingly, a minimal model for the mutual coupling between redox energy and acid-base energy through an electronically energized and protonated ferricytochrome b-559 intermediate is proposed. FCCP would exert its uncoupling action by removing the proton from the energized oxidized form of the cytochrome high-potential couple. Conversely, protonation of the basal oxidized form of the low-potential couple-either directly at low pH or indirectly at high pH through redox reactions—would cause its energization.