Abstract
One of the key unresolved issues regarding proton pumping in cytochrome c oxidase (CcO) is the identity of the gating element that prevents the backflow of protons. In this study, we analyze two popular proposals for this element: isomerization of the key branching residue (Glu-286) and (re)orientation of water molecules in the hydrophobic cavity. Using a multifaceted set of computational analyses that involve CcO embedded in either an implicit or explicit treatment of lipid membrane, we show that neither Glu-286 nor active-site water likely constitutes the gating element. Detailed energetic and structural analyses of the simulation results indicate that the gating-relevant properties of these structural motifs observed in previous work are likely a result of the simplified computational models employed in those studies.