Oxidation-reduction potential measurements of cytochrome c peroxidase and pH dependent spectral transitions in the ferrous enzyme.
The redox potential of the ferrous/ferric couple in cytochrome c peroxidase has been measured as a function of pH between pH 4.5 and 8. The redox potential decreases linearly as a function of pH between pH 4.5 and 7 with a slope of --57 +/- 2 mV per pH unit. Above pH 7, there is a positive inflection in the midpoint potential versus pH plot attributed to an ionizable group in the ferrous enzyme with pKa of 7.6 +/- 0.1. The midpoint potential at pH 7 is--0.194 V relative to the standard hydrogen electrode at 25 degree C. Ferrocytochrome c peroxidase undergoes a reversible spectral transition as a function of pH. Below pH 7, the enzyme has a spectrum typical of high spin ferroheme proteins while above pH 8, the spectrum is typical of low spin ferroheme proteins. The transition is caused by a co-operative, two proton ionization with an apparent pKa of 7.7 +/- 0.2. Two other single proton ionizations cause minor perturbations to the spectrum of ferrocytochrome c peroxidase. One has a pKa of 5.7 +/- 0.2 while the second has a pKa of 9.4 +/- 0.2.
D003578:Cytochrome-c Peroxidase; D006863:Hydrogen-Ion Concentration; D007501:Iron; D010084:Oxidation-Reduction; D010544:Peroxidases; D011199:Potentiometry; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D013053:Spectrophotometry; D013056:Spectrophotometry, Ultraviolet
D007501:Iron; D010544:Peroxidases; D003578:Cytochrome-c Peroxidase