Control Mechanisms of Electron and Proton Transfer in Cytochrome c Oxidase
Cytochrome c oxidase is a remarkable molecular machine that is bound in the inner mitochondrial membrane or in the cell membrane of bacteria. Using the free energy that is released when electrons from foodstuff are transferred to molecular oxygen, protons are translocated from the inner to the outer side of the membrane. In addition, reduction of each dioxygen molecule to water is associated with the uptake of four substrate protons from the inner side of the membrane. These reactions add to the generation of an electro-chemical gradient that is subsequently used for the synthesis of the mobile energy carrier, ATP.
In order for this proton-pumping machinery to work efficiently, the transfer of protons and electrons through the enzyme must be carefully regulated. The focus of this thesis is to elucidate the coupling mechanisms between electron and proton transfers. The temporal sequence of intimately coupled intermediate states, that have previously not been observed, has been deduced and a common mechanism for electron transfer to the binuclear centre has been formulated. In this mechanism, electron transfer to the binuclear centre is thermodynamically unfavourable but proton transfer from adjacent protonatable groups traps the electron.
The conclusions are based mainly on analysis of data obtained from time-resolved optical-absorption measurements on enzyme isolated from R. sphae-roides or bovine heart. One important tool in the investigations has been the use of deuterium oxide as the reaction solvent. Other approaches include the use of site-directed mutants, designed to specifically alter the rates and thermo-dynamics of electron and proton transfer to oxygen.