Proton pumping mechanism and catalytic cycle of cytochrome c oxidase: Coulomb pump model with kinetic gating

122

146

Aerobic respiration in bacteria, Archaea, and mitochondria is performed by oxygen reductase members of the heme-copper oxidoreductase superfamily. These enzymes are redox-driven proton pumps which conserve part of the free energy released from oxygen reduction to generate a proton motive force. The oxygen reductases can be divided into three main families based on evolutionary and structural analyses (A-, B- and C-families), with the B- and C-families evolving after the A-family. The A-family utilizes two proton input channels to transfer protons for pumping and chemistry, whereas the B- and C-families require only one. Generally, the B- and C-families also have higher apparent oxygen affinities than the A-family. Here we use whole cell proton pumping measurements to demonstrate differential proton pumping efficiencies between representatives of the A-, B-, and C-oxygen reductase families. The A-family has a coupling stoichiometry of 1 H + /e - , whereas the B- and C-families have coupling stoichiometries of 0.5 H + /e - . The differential proton pumping stoichiometries, along with differences in the structures of the proton-conducting channels, place critical constraints on models of the mechanism of proton pumping. Most significantly, it is proposed that the adaptation of aerobic respiration to low oxygen environments resulted in a concomitant reduction in energy conservation efficiency, with important physiological and ecological consequences.

Section: Discussionmentioning

confidence: 99%

Adaptation of aerobic respiration to low O ₂ environments

Han

Hemp

Pace

et al. 2011

122

146

“…The D pathway terminates at a highly conserved glutamic acid residue in subunit I, E242 (numbering based on the bovine heart enzyme). From here the protons to be pumped across the membrane are suggested first to be carried to a pump site (also called a protonloading site, PLS) in a redox-dependent manner (6)(7)(8)(9)(10). The proton at the PLS is generally agreed to be ejected to the positively charged P side of the membrane by electrostatic repulsion from uptake of a chemical proton into the BNC, as suggested independently by Morgan et al (11) and Rich (12).…”

mentioning

confidence: 84%

Computational study of the activated O _H state in the catalytic mechanism of cytochrome c oxidase

Sharma

Karlin

Wikström

2013

119

Complex IV in the respiratory chain of mitochondria and bacteria catalyzes reduction of molecular oxygen to water, and conserves much of the liberated free energy as an electrochemical proton gradient, which is used for the synthesis of ATP. Photochemical electron injection experiments have shown that reduction of the ferric/cupric state of the enzyme's binuclear heme a 3 /Cu B center is coupled to proton pumping across the membrane, but only if oxidation of the reduced enzyme by O 2 immediately precedes electron injection. In contrast, reduction of the binuclear center in the "as-isolated" ferric/cupric enzyme is sluggish and without linkage to proton translocation. During turnover, the binuclear center apparently shuttles via a metastable but activated ferric/cupric state (O H ), which may decay into a more stable catalytically incompetent form (O) in the absence of electron donors. The structural basis for the difference between these two states has remained elusive, and is addressed here using computational methodology. The results support the notion that Cu B [II] is either three-coordinated in the O H state or shares an OH − ligand with heme a 3 in a strained μ-hydroxo structure. Relaxation to state O is initiated by hydration of the binuclear site. The redox potential of Cu B is expected, and found by density functional theory calculations, to be substantially higher in the O H state than in state O. Our calculations also suggest that the neutral radical form of the cross-linked tyrosine in the binuclear site may be more significant in the catalytic cycle than suspected so far.oxygen reduction | electron transfer

“…S12) as it requires that proton transfer to the pumping site precede that to the BNC, a feature that would minimize the amount of "slippage," where chemistry is done without pumping. The model also provides a microscopic framework for the kinetic gating phenomena identified in the kinetic network analysis (50) and discussed previously (51,52). Although our calculations focus on states implicated in the P R to ′F transition, because the key driving force for the Glu286 pK′ 7 modulation is protonation of the presumed pumping site (PRDa 3 ), it does not depend on the specific chemical state of the BNC.…”

Section: Microscopic Qm/mm-ti Pk′ 7 Calculations and The Effect Of Cmentioning

confidence: 99%

Changing hydration level in an internal cavity modulates the proton affinity of a key glutamate in cytochromecoxidase

Goyal

Yang

et al. 2013

112

Cytochrome c oxidase contributes to the transmembrane proton gradient by removing two protons from the high-pH side of the membrane each time the binuclear center active site is reduced. One proton goes to the binuclear center, whereas the other is pumped to the low-pH periplasmic space. Glutamate 286 (Glu286) has been proposed to serve as a transiently deprotonated proton donor. Using unrestrained atomistic molecular dynamics simulations, we show that the size of and water distribution in the hydrophobic cavity that holds Glu286 is controlled by the protonation state of the propionic acid of heme a 3 , a group on the proton outlet pathway. Protonation of the propionate disrupts hydrogen bonding to two side chains, allowing a loop to swing open. Continuum electrostatics and atomistic free-energy perturbation calculations show that the resultant changes in hydration and electrostatic interactions lower the Glu proton affinity by at least 5 kcal/mol. These changes in the internal hydration level occur in the absence of major conformational transitions and serve to stabilize needed transient intermediates in proton transport. The trigger is not the protonation of the Glu of interest, but rather the protonation of a residue ∼10 Å away. Thus, unlike local water penetration to stabilize a new charge, this finding represents a specific role for water molecules in the protein interior, mediating proton transfers and facilitating ion transport.proton pumping | pK a W ater is essential to the structure, dynamics, and function of biomolecules (1, 2), and its role in protein folding, association (3), and dynamics (4, 5) has been well documented. The highly polar and polarizable water molecules play diverse roles in protein interiors. Water can aid catalysis in enzyme active sites (6-8). Water or water chains are often observed in proteins that are (9, 10) proton or ion transporters or pumps (11)(12)(13)(14). Internal cavities holding functional water molecules are believed to have a fairly constant level of hydration throughout the protein reaction cycle, unless significant conformational changes occur (15). Water penetration in response to the ionization or reduction of internal groups has been extensively discussed (16,17), although it is usually described as part of protein's local dielectric response.Cytochrome c oxidase (CcO) adds to the transmembrane proton gradient through proton transport coupled to electron transfer reactions (12,18,19). In the overall reaction, electrons from four cytochromes c are transferred to oxygen to make two water molecules at the binuclear center (BNC). The four protons needed for chemistry are bound only from the high-pH, N side of the membrane. Coupled to the process, four more protons are transferred across the membrane from the high-to low-pH (P) side of the membrane. Thus, eight charges are transferred across the membrane as each O 2 is reduced.Glu286 is a required, conserved residue that is expected to transfer protons from the D channel either to the BNC or the proton-loading site (...