Structural Changes and Proton Transfer in Cytochrome c Oxidase

Proton pumping A-type cytochrome oxidase (CO) terminates the respiratory chains of mitochondria and many bacteria. Three possible proton transfer pathways (D, K, and H channels) have been identified based on structural, functional, and mutational data. Whereas the D channel provides the route for all pumped protons in bacterial A-type COs, studies of bovine mitochondrial CO have led to suggestions that its H channel instead provides this route. Here, we have studied H-channel function by performing atomistic molecular dynamics simulations on the entire, as well as core, structure of bovine CO in a lipid-solvent environment. The majority of residues in the H channel do not undergo large conformational fluctuations. Its upper and middle regions have adequate hydration and H-bonding residues to form potential proton-conducting channels, and Asp51 exhibits conformational fluctuations that have been observed crystallographically. In contrast, throughout the simulations, we do not observe transient water networks that could support proton transfer from the N phase toward heme via neutral His413, regardless of a labile H bond between Ser382 and the hydroxyethylfarnesyl group of heme In fact, the region around His413 only became sufficiently hydrated when His413 was fixed in its protonated imidazolium state, but its calculated pK is too low for this to provide the means to create a proton transfer pathway. Our simulations show that the electric dipole moment of residues around heme changes with the redox state, hence suggesting that the H channel could play a more general role as a dielectric well.

Section: Discussionmentioning

confidence: 55%

Section: Model System Cu a Heme A Bnc His413 Asp51 Glu242 Tyr244 Lys3mentioning

confidence: 82%

Insights into functions of the H channel of cytochrome c oxidase from atomistic molecular dynamics simulations

Sharma

Jambrina

Kaukonen

et al. 2017

“…It would also be interesting to address the role of the Rcf proteins in controlling the ratio of O 2 reduction over ATP formation (the P/O ratio) because changes in structure, such as those changes presumably induced by removal of Rcf1, may interfere with proton pumping (49)(50)(51)(52).…”

Section: Discussionmentioning

confidence: 99%

Regulatory role of the respiratory supercomplex factors in Saccharomyces cerevisiae

Lundin

Ballmoos

Ott

et al. 2016

Self Cite

The respiratory supercomplex factors (Rcf) 1 and 2 mediate supramolecular interactions between mitochondrial complexes III (ubiquinolcytochrome c reductase; cyt. bc 1 ) and IV (cytochrome c oxidase; CytcO). In addition, removal of these polypeptides results in decreased activity of CytcO, but not of cyt. bc 1 . In the present study, we have investigated the kinetics of ligand binding, the singleturnover reaction of CytcO with O 2 , and the linked cyt. bc 1 -CytcO quinol oxidation-oxygen-reduction activities in mitochondria in which Rcf1 or Rcf2 were removed genetically (strains rcf1Δ and rcf2Δ, respectively). The data show that in the rcf1Δ and rcf2Δ strains, in a significant fraction of the population, ligand binding occurs over a time scale that is ∼100-fold faster (τ ≅ 100 μs) than observed with the wild-type mitochondria (τ ≅ 10 ms), indicating structural changes. This effect is specific to removal of Rcf and not dissociation of the cyt. bc 1 -CytcO supercomplex. Furthermore, in the rcf1Δ and rcf2Δ strains, the single-turnover reaction of CytcO with O 2 was incomplete. This observation indicates that the lower activity of CytcO is caused by a fraction of inactive CytcO rather than decreased CytcO activity of the entire population. Furthermore, the data suggest that the Rcf1 polypeptide mediates formation of an electrontransfer bridge from cyt. bc 1 to CytcO via a tightly bound cyt. c. We discuss the significance of the proposed regulatory mechanism of Rcf1 and Rcf2 in the context of supramolecular interactions between cyt. bc 1 and CytcO.cytochrome c oxidase | electron transfer | membrane protein | cytochrome aa 3 | cytochrome bc 1

“…To interrogate the effect of S425 in RsCcO (equivalent to S382 in bCcO) on proton translocation in the D pathway, Brzezinski and coworkers (42) mutated S425 to alanine and measured its proton translocation efficiency during the O 2 reaction cycle as a function of pH. They found that structural changes in S425 propagate to the D pathway, thereby modulating proton transfer rates (42). These data suggest that the S382-linked allosteric structural transition may play a role in proton translocation in both mammalian and bacterial oxidases.…”

Section: Discussionmentioning

confidence: 99%

Crystal structure of CO-bound cytochrome c oxidase determined by serial femtosecond X-ray crystallography at room temperature

Ishigami

Zatsepin

Hikita

et al. 2017

Cytochrome c oxidase (CcO), the terminal enzyme in the electron transfer chain, translocates protons across the inner mitochondrial membrane by harnessing the free energy generated by the reduction of oxygen to water. Several redox-coupled proton translocation mechanisms have been proposed, but they lack confirmation, in part from the absence of reliable structural information due to radiation damage artifacts caused by the intense synchrotron radiation. Here we report the room temperature, neutral pH (6.8), damage-free structure of bovine CcO (bCcO) in the carbon monoxide (CO)-bound state at a resolution of 2.3 Å, obtained by serial femtosecond X-ray crystallography (SFX) with an X-ray free electron laser. As a comparison, an equivalent structure was obtained at a resolution of 1.95 Å, from data collected at a synchrotron light source. In the SFX structure, the CO is coordinated to the heme a 3 iron atom, with a bent Fe-C-O angle of ∼142°. In contrast, in the synchrotron structure, the Fe-CO bond is cleaved; CO relocates to a new site near Cu B , which, in turn, moves closer to the heme a 3 iron by ∼0.38 Å. Structural comparison reveals that ligand binding to the heme a 3 iron in the SFX structure is associated with an allosteric structural transition, involving partial unwinding of the helix-X between heme a and a 3 , thereby establishing a communication linkage between the two heme groups, setting the stage for proton translocation during the ensuing redox chemistry.bioenergetics | X-ray free electron laser | crystallography | cytochrome c oxidase | serial femtosecond crystallography