G. Battistuzzi et al., REDOX THERMODYNAMICS OF THE NATIVE AND ALKALINE FORMS OF EUKARYOTIC AND BACTERIAL CLASS-I CYTOCHROMES-C, Biochemistry, 36(51), 1997, pp. 16247-16258
The reduction potentials of beef heart cytochrome c and cytochromes c(
2) from Rhodopseudomonas palustris, Rhodobacter sphaeroides, and Rhodo
bacter capsulatus were measured through direct electrochemistry at a s
urface-modified gold electrode as a function of temperature in nonisot
hermal experiments carried out at neutral and alkaline pH values. The
thermodynamic parameters for protein reduction (Delta S(rc)degrees and
Delta H(rc)degrees) were determined for the native and alkaline confo
rmers. Enthalpy and entropy terms underlying species-dependent differe
nces in E degrees and pH-and temperature-induced E degrees changes for
a given cytochrome were analyzed. The difference of about +0.1 V in E
degrees between cytochromes c(2) and the eukaryotic species can be se
parated into an enthalpic term (-Delta Delta H(rc)degrees/F) of +0.130
V and an entropic term (T Delta Delta S(rc)degrees/F) of -0.040 V. He
nce, the higher potential of the bacterial species appears to be deter
mined entirely by a greater enthalpic stabilization of the reduced sta
te. Analogously, the much lower potential of the alkaline conformer(s)
as compared to the native species is by far enthalpic in origin for b
oth protein families, and is largely determined by the substitution of
Met for Lys in axial heme ligation. Instead, the biphasic E degrees/t
emperature profile for the native cytochromes is due to a difference i
n reduction entropy between the conformers at low and high temperature
s. Temperature-dependent H-1 NMR experiments suggest that the temperat
ure-induced transition also involves a change in orientation of the ax
ial methionine ligand with respect to the heme plane.