Involvement of a labile axial histidine in coupling electron and proton transfer in Methylophilus methylotrophus cytochrome c''.
Methylophilus methylotrophus cytochrome c'' is an unusual monohaem protein (15 kDa) undergoing a redox-linked spin-state transition [Santos, H. & Turner, D. L. (1988) Biochim. Biophys. Acta 954, 277-286]. The midpoint redox potential of cytochrome c" was measured over the pH range 4-10. The pH dependence of the midpoint redox potential was interpreted in terms of a model that considers the redox-state dependence of the ionization of two distinct and non-interacting protonated groups in the protein. This analysis led to the following pKa values within the pH range studied: pKa10 = 6.4, pKa1r = 5.4 and pKa2r = 8.1. Proton-NMR spectroscopy was used to assist the characterization of the two ionizing groups responsible for the observed redox-Bohr effect: the group ionizing with a lower pKar was assigned to a haem propionic acid substituent and the other to the axial histidine ligand which becomes detached upon reduction, which has a pKa0 too low to be measured. It is shown that M. methylotrophus cytochrome c" is able to couple electron and proton transfer in the physiological pH range through a mechanism involving reversible change in the haem-iron coordination. Possible implications for the physiological role of the protein are discussed.
Costa
H S
HS
Centro de Tecnologia Química e Biológica, Oeiras, Portugal.
Santos
H
H
Turner
D L
DL
Xavier
A V
AV
D000596:Amino Acids / Q000032:analysis; D003574:Cytochrome c Group / Q000737:chemistry* / Q000378:metabolism; D004579:Electron Transport; D006088:Gram-Negative Aerobic Bacteria / Q000737:chemistry*; D006639:Histidine / Q000737:chemistry* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy; D010084:Oxidation-Reduction; D011522:Protons
D000596:Amino Acids; D003574:Cytochrome c Group; D011522:Protons; C076615:cytochrome c''; D006639:Histidine