Redox chemistry and acid-base equilibria of mitochondrial plant cytochromes c

Mitochondrial cytochromes c from spinach, cucumber, and sweet potato have b een investigated through direct electrochemical measurements and electronic and H-1 NMR spectroscopies, under conditions of varying temperature and pH . The solution behaviors of these plant cytochromes closely resemble, but d o not fully reproduce, those of homologous eukaryotic species. The reductio n potentials (E degrees') at pH 7 and 25 degrees C are +0.268 V (spinach), +0.271 V (cucumber), and +0.274 V (sweet potato) vs SHE. Three acid-base eq uilibria have been determined for the oxidized proteins with apparent pK(a) values of 2.5, 4.8, and 8.3-8.9, which are related to disruption of axial heme ligation, deprotonation of the solvent-exposed heme propionate-7 and r eplacement of the methionine axially bound to the heme iron with a stronger ligand, respectively. The most significant peculiarities with respect to t he mammalian analogues include: (i) less negative reduction enthalpies and entropies (Delta S degrees'(rc) and Delta H degrees'(rc)) for the various p rotein conformers [low- and high-T native (N-1 and N-2) and alkaline (A)], whose effects at pH 7 and 25 degrees C largely compensate to produce E degr ees' values very similar to those of the mammalian proteins; (ii) the N-1 - N-2 transition that occurs at a lower temperature (e.g., 30-35 degrees C v s 50 degrees C at pH 7.5) and at a lower pH (7 vs 7.5); and (iii) a more pr onounced temperature-induced decrease in the pK(a) for the alkaline transit ion which allows observation of the alkaline conformer(s) at pH values as l ow as 7 upon increasing the temperature above 40 degrees C. Regarding the p H and the temperature ranges of existence of the various protein conformers , these plant cytochromes c are closer to bacterial cytochromes c(2).