Water exit pathways and proton pumping mechanism in B-type cytochrome c oxidase from molecular dynamics simulations

• Published in: Biochimica et biophysica acta 2016-09, Vol.1857 (9), p.1594-1606
• Main Authors: Yang, Longhua, Skjevik, Åge A., Han Du, Wen-Ge, Noodleman, Louis, Walker, Ross C., Götz, Andreas W.
• Format: Article
• Language: English
• Subjects: Computer simulation; Cytochrome c oxidase; Electron Transport Complex IV - chemistry; Molecular dynamics; Molecular Dynamics Simulation; Proton pumping mechanism; Proton Pumps; Proton transport; Protons; Water - chemistry; Water exit pathway
• ISSN: 0005-2728; 0006-3002; 1879-2650
• DOI: 10.1016/j.bbabio.2016.06.005

Cytochrome c oxidase (CcO) is a vital enzyme that catalyzes the reduction of molecular oxygen to water and pumps protons across mitochondrial and bacterial membranes. While proton uptake channels as well as water exit channels have been identified for A-type CcOs, the means by which water and protons exit B-type CcOs remain unclear. In this work, we investigate potential mechanisms for proton transport above the dinuclear center (DNC) in ba3-type CcO of Thermus thermophilus. Using long-time scale, all-atom molecular dynamics (MD) simulations for several relevant protonation states, we identify a potential mechanism for proton transport that involves propionate A of the active site heme a3 and residues Asp372, His376 and Glu126II, with residue His376 acting as the proton-loading site. The proposed proton transport process involves a rotation of residue His376 and is in line with experimental findings. We also demonstrate how the strength of the salt bridge between residues Arg225 and Asp287 depends on the protonation state and that this salt bridge is unlikely to act as a simple electrostatic gate that prevents proton backflow. We identify two water exit pathways that connect the water pool above the DNC to the outer P-side of the membrane, which can potentially also act as proton exit transport pathways. Importantly, these water exit pathways can be blocked by narrowing the entrance channel between residues Gln151II and Arg449/Arg450 or by obstructing the entrance through a conformational change of residue Tyr136, respectively, both of which seem to be affected by protonation of residue His376. [Display omitted] •We performed long-time scale molecular dynamics simulations on ba3-type cytochrome c oxidase from Thermus thermophilus.•A mechanism for proton transport above the dinuclear center (DNC) is proposed, with residue His376 as proton-loading site.•The salt bridge Arg225–Asp287 is stable in several protonation states and unlikely to act as a simple electrostatic gate.•Two water exit pathways that connect the water pool above the DNC to the outer P-side of the membrane are identified.•Protonation of His376 blocks both water pathways by narrowing the entrance channel or conformational changes of Tyr136.