THE COUPLING OF ELECTRON-TRANSFER AND PROTON TRANSLOCATION - ELECTROSTATIC CALCULATIONS ON PARACOCCUS-DENITRIFICANS CYTOCHROME-C-OXIDASE

We have calculated the electrostatic potential and interaction energie s of ionizable groups and analyzed the response of the protein environ ment to redox changes in Paracoccus denitrificans cytochrome c oxidase by using a continuum dielectric model and finite difference technique . Subsequent Monte Carlo sampling of protonation states enabled us to calculate the titration curves of all protonatable groups in the enzym e complex. Inclusion of a model membrane allowed us to restrict the ca lculations to the functionally essential subunits I and II. Some resid ues were calculated to have complex titration curves, as a result of s trong electrostatic coupling, desolvation, and dipolar interactions. A round the heme a(3)-Cu-B binuclear center, we have identified a cluste r of 18 strongly interacting residues that account for most of the pro ton uptake linked to electron transfer. This was calculated to be betw een 0.7 and 1.1 H+ per electron, depending on the redox transition con sidered. A hydroxide ion bound to Cu, was determined to become protona ted to form water upon transfer of the first electron to the binuclear site. The bulk of the protonation changes linked to further reduction of the hems a(3)-Cu-B center was calculated to be due to proton uptak e by the interacting cluster and Glu(II-78) Upon formation of the thre e-electron reduced state (P1), His(325), modeled in an alternative ori entation away from Cu-B, was determined to become protonated. The agre ement of these results with experiment and their relevance in the ligh t of possible mechanisms of redox-coupled proton transfer are discusse d.