Cholate Disrupts Regulatory Functions of Cytochrome c Oxidase.

Cytochrome c oxidase (CytOx), the oxygen-accepting and rate-limiting enzyme of mitochondrial respiration, binds with 10 molecules of ADP, 7 of which are exchanged by ATP at high ATP/ADP-ratios. These bound ATP and ADP can be exchanged by cholate, which is generally used for the purification of CytOx. Many crystal structures of isolated CytOx were performed with the enzyme isolated from mitochondria using sodium cholate as a detergent.

Multiple Mechanisms Regulate Eukaryotic Cytochrome C Oxidase.

Cytochrome c oxidase (COX), the rate-limiting enzyme of mitochondrial respiration, is regulated by various mechanisms. Its regulation by ATP (adenosine triphosphate) appears of particular importance, since it evolved early during evolution and is still found in cyanobacteria, but not in other bacteria. Therefore the "allosteric ATP inhibition of COX" is described here in more detail.

Complex IV - The regulatory center of mitochondrial oxidative phosphorylation.

ATP, the universal energy currency in all living cells, is mainly synthesized in mitochondria by oxidative phosphorylation (OXPHOS). The final and rate limiting step of the respiratory chain is cytochrome c oxidase (COX) which represents the regulatory center of OXPHOS. COX is regulated through binding of various effectors to its "supernumerary" subunits, by reversible phosphorylation, and by expression of subunit isoforms.

Regulation of cytochrome c oxidase contributes to health and optimal life.

The generation of cellular energy in the form of ATP occurs mainly in mitochondria by oxidative phosphorylation. Cytochrome c oxidase (CytOx), the oxygen accepting and rate-limiting step of the respiratory chain, regulates the supply of variable ATP demands in cells by "allosteric ATP-inhibition of CytOx." This mechanism is based on inhibition of oxygen uptake of CytOx at high ATP/ADP ratios and low ferrocytochrome c concentrations in the mitochondrial matrix cooperative interaction of the two substrate binding sites in dimeric CytOx.

Stress-mediated generation of deleterious ROS in healthy individuals - role of cytochrome c oxidase.

Psychosocial stress is known to cause an increased incidence of coronary heart disease. In addition, multiple other diseases like cancer and diabetes mellitus have been related to stress and are mainly based on excessive formation of reactive oxygen species (ROS) in mitochondria. The molecular interactions between stress and ROS, however, are still unknown.

Reversible dimerization of cytochrome c oxidase regulates mitochondrial respiration.

Almost all energy consumed by higher organisms, either in the form of ATP or heat, is produced in mitochondria by respiration and oxidative phosphorylation through five protein complexes in the inner membrane. High-resolution x-ray analysis of crystallized cytochrome c oxidase (CytOx), the final oxygen-accepting complex of the respiratory chain, isolated by using cholate as detergent, revealed a dimeric structure with 13 subunits in each monomer. In contrast, CytOx isolated with non-ionic detergents appeared to be monomeric.

Regulation of Mammalian 13-Subunit Cytochrome c Oxidase and Binding of other Proteins: Role of NDUFA4.

Cytochrome c oxidase (CcO) is the final oxygen accepting enzyme complex (complex IV) of the mitochondrial respiratory chain. In contrast to the other complexes (I, II, and III), CcO is highly regulated via isoforms for six of its ten nuclear-coded subunits, which are differentially expressed in species, tissues, developmental stages, and cellular oxygen concentrations. Recent publications have claimed that NADH dehydrogenase (ubiquinone) 1 alpha subcomplex 4 (NDUFA4), originally identified as subunit of complex I, represents a 14th subunit of CcO.

The subunit composition and function of mammalian cytochrome c oxidase.

Cytochrome c oxidase (COX) from mammals and birds is composed of 13 subunits. The three catalytic subunits I-III are encoded by mitochondrial DNA, the ten nuclear-coded subunits (IV, Va, Vb, VIa, VIb, VIc, VIIa, VIIb, VIIc, VIII) by nuclear DNA. The nuclear-coded subunits are essentially involved in the regulation of oxygen consumption and proton translocation by COX, since their removal or modification changes the activity and their mutation causes mitochondrial diseases.

High efficiency versus maximal performance--the cause of oxidative stress in eukaryotes: a hypothesis.

Degenerative diseases are in part based on elevated production of ROS (reactive oxygen species) in mitochondria, mainly during stress and excessive work under stress (strenuous exercise). The production of ROS increases with increasing mitochondrial membrane potential (ΔΨ(m)). A mechanism is described which is suggested to keep ΔΨ(m) at low values under normal conditions thus preventing ROS formation, but is switched off under stress and excessive work to maximize the rate of ATP synthesis, accompanied by decreased efficiency.

Individual biochemical behaviour versus biological robustness: spotlight on the regulation of cytochrome c oxidase.

During evolution from prokaryotes to eukaryotes, the main function of cytochrome c oxidase (COX), i.e., the coupling of oxygen reduction to proton translocation without the production of ROS (reactive oxygen species) remained unchanged demonstrating its robustness.

Introduction to mitochondrial oxidative phosphorylation.

The basic mechanism of ATP synthesis in the mitochondria by oxidative phosphorylation (OxPhos) was revealed in the second half of the twentieth century. The OxPhos complexes I-V have been analyzed concerning their subunit composition, genes, and X-ray structures. This book presents new developments regarding the morphology, biogenesis, gene evolution, heat, and reactive oxygen species (ROS) generation in mitochondria, as well as the structure and supercomplex formation of OxPhos complexes.

Discovering the phosphoproteome of the hydrophobic cytochrome c oxidase membrane protein complex.

Many cellular processes are regulated by reversible phosphorylation to change the activity state of proteins. One example is cytochrome c oxidase (COX) with its important function for energy metabolism in the mitochondria. The phosphorylation of this enzyme is a prerequisite for the allosteric ATP-inhibition and therefore necessary to adapt energy production to ATP demand of the cell.

Multiple phosphorylations of cytochrome c oxidase and their functions.

Cytochrome c oxidase (COX), the terminal enzyme of the mitochondrial electron transport chain, is regulated by isozyme expression, allosteric effectors such as the ATP/ADP ratio, and reversible phosphorylation. Of particular interest is the "allosteric ATP-inhibition," which has been hypothesized to keep the mitochondrial membrane potential at low healthy values (<140 mV), thus preventing the formation of superoxide radical anions, which have been implicated in multiple degenerative diseases. It has been proposed that the "allosteric ATP-inhibition" is switched on by the protein kinase A-dependent phosphorylation of COX.

The role of mitochondrial membrane potential in ischemic heart failure.

The molecular events occurring during myocardial infarction and cardioprotection are described with an emphasis on the changes of the mitochondrial membrane potential (ΔΨ(m)). The low ΔΨ(m) values of the normal beating heart (100-140 mV) are explained by the allosteric ATP-inhibition of cytochrome c oxidase (CcO) through feedback inhibition by ATP at high [ATP]/[ADP] ratios. During ischemia the mechanism is reversibly switched off by signaling through reactive oxygen species (ROS).

Mitochondrial respiration and membrane potential are regulated by the allosteric ATP-inhibition of cytochrome c oxidase.

This paper describes the problems of measuring the allosteric ATP-inhibition of cytochrome c oxidase (CcO) in isolated mitochondria. Only by using the ATP-regenerating system phosphoenolpyruvate and pyruvate kinase full ATP-inhibition of CcO could be demonstrated by kinetic measurements. The mechanism was proposed to keep the mitochondrial membrane potential (DeltaPsi(m)) in living cells and tissues at low values (100-140 mV), when the matrix ATP/ADP ratios are high.

New extension of the Mitchell Theory for oxidative phosphorylation in mitochondria of living organisms.

The Mitchell Theory implies the proton motive force Deltap across the inner mitochondrial membrane as the energy-rich intermediate of oxidative phosphorylation. Deltap is composed mainly of an electrical (DeltaPsi(m)) and a chemical part (DeltapH) and generated by the respiratory chain complexes I, III and IV. It is consumed mostly by the ATP synthase (complex V) to produce ATP.

Degenerative diseases, oxidative stress and cytochrome c oxidase function.

Aging and degenerative diseases are associated with increased levels of reactive oxygen species (ROS). ROS are mostly produced in mitochondria, and their levels increase with higher mitochondrial membrane potential. Cellular respiratory control is based on inhibition of respiration by high membrane potentials.

Phosphorylation and kinetics of mammalian cytochrome c oxidase.

The influence of protein phosphorylation on the kinetics of cytochrome c oxidase was investigated by applying Western blotting, mass spectrometry, and kinetic measurements with an oxygen electrode. The isolated enzyme from bovine heart exhibited serine, threonine, and/or tyrosine phosphorylation in various subunits, except subunit I, by using phosphoamino acid-specific antibodies. The kinetics revealed slight inhibition of oxygen uptake in the presence of ATP, as compared with the presence of ADP.

The possible role of cytochrome c oxidase in stress-induced apoptosis and degenerative diseases.

Apoptotic cell death can occur by two different pathways. Type 1 is initiated by the activation of death receptors (Fas, TNF-receptor-family) on the plasma membrane followed by activation of caspase 8. Type 2 involves changes in mitochondrial integrity initiated by various effectors like Ca(2+), reactive oxygen species (ROS), Bax, or ceramide, leading to the release of cytochrome c and activation of caspase 9.

Intrinsic and extrinsic uncoupling of oxidative phosphorylation.

This article reviews parameters of extrinsic uncoupling of oxidative phosphorylation (OxPhos) in mitochondria, based on induction of a proton leak across the inner membrane. The effects of classical uncouplers, fatty acids, uncoupling proteins (UCP1-UCP5) and thyroid hormones on the efficiency of OxPhos are described. Furthermore, the present knowledge on intrinsic uncoupling of cytochrome c oxidase (decrease of H(+)/e(-) stoichiometry=slip) is reviewed.

Control of mitochondrial membrane potential and ROS formation by reversible phosphorylation of cytochrome c oxidase.

Phosphorylation of isolated cytochrome c oxidase from bovine kidney and heart, and of the reconstituted heart enzyme, with protein kinase A, cAMP and ATP turns on the allosteric ATP-inhibition at high ATP/ADP ratios. Also incubation of isolated bovine liver mitochondria only with cAMP andATP turns on, and subsequent incubation with Ca2+ turns off the allosteric ATP-inhibition of cytochrome c oxidase. In the bovine heart enzyme occur only three consensus sequences for cAMP-dependent phosphorylation (in subunits I, III and Vb).