Dataset Information

Effects of inhibitory ligands on the aerobic carbon monoxide complex of cytochrome c oxidase.

ABSTRACT: 1. In the presence of both CO and O2, ox heart cytochrome c oxidase forms a 607 nm-peak intermediate distinct from both the cytochrome a2+a3 2+CO and the cytochrome a3+a3 2+CO ('mixed-valence') CO complexes. 2. This aerobic CO compound is stable towards ferricyanide addition, but decomposed on treatment with ferric cytochrome a2 ligands such as formate, cyanide and azide. 3. Addition of formate or cyanves rise to a complex with alpha-peak at 598 nm, not identical with any azide complex of the free enzyme, but possibly a cytochrome a3 2+NO complex produced by oxidative attack of partially reduced O2 on the azide. 4. The results support the idea that although the initial reaction of oxygen is with cytochrome a3 2+, the next step is not an oxidation of the ferrous cytochrome a3, but a transfer of O2 to a neighbouring group, such as Cu+, to give Cu2+O2- or similar complexes. 5. The aerobic CO complex is then identified as a3+a3 2+COCu2+O2-; a similar compound ('Compound C') is formed by photolysis of a3+a3 2+CO (the 'mixed-valence' CO complex) in the presence of oxygen at low temperatures.

SUBMITTER: Nicholls P

PROVIDER: S-EPMC1161632 | biostudies-other | 1979 Dec

REPOSITORIES: biostudies-other

ACCESS DATA

Similar Datasets

Project description:Commonly used anesthetics induce widespread neuronal degeneration in the developing mammalian brain via the oxidative-stress-associated mitochondrial apoptosis pathway. Dysregulation of cytochrome oxidase (CcOX), the terminal oxidase of the electron transport chain, can result in reactive oxygen species (ROS) formation. Isoflurane has previously been shown to activate this enzyme. Carbon monoxide (CO), as a modulator of CcOX, is of interest because infants and children are routinely exposed to CO during low-flow anesthesia. We have recently demonstrated that low concentrations of CO limit and prevent isoflurane-induced neurotoxicity in the forebrains of newborn mice in a dose-dependent manner. However, the effect of CO on CcOX in the context of anesthetic-induced oxidative stress is unknown. Seven-day-old male CD-1 mice underwent 1h exposure to 0 (air), 5, or 100ppm CO in air with or without isoflurane. Exposure to isoflurane or CO independently increased CcOX kinetic activity and increased ROS within forebrain mitochondria. However, exposure to CO combined with isoflurane paradoxically limited CcOX activation and oxidative stress. There were no changes seen in steady-state levels of CcOX I protein, indicating post-translational modification of CcOX as an etiology for changes in enzyme activity. CO exposure led to differential effects on CcOX subunit I tyrosine phosphorylation depending on concentration, while combined exposure to isoflurane with CO markedly increased the enzyme phosphorylation state. Phosphorylation of tyrosine 304 of CcOX subunit I has been shown to result in strong enzyme inhibition, and the relative reduction in CcOX kinetics following exposure to CO combined with isoflurane may have been due, in part, to such phosphorylation. Taken together, the data suggest that CO modulates CcOX in the developing brain during isoflurane exposure, thereby limiting oxidative stress. These CO-mediated effects could have implications for the development of low-flow anesthesia in infants and children to prevent anesthesia-induced oxidative stress.

Project description:Inner membranes were prepared from Escherichia coli strain RG 145, which is deficient in cytochrome bd, but overexpresses cytochrome bo [Au and Gennis (1987) J. Bacteriol. 169, 3237-3242]. The latter was purified 7-fold by extracting the membranes with octyl beta-D-glucopyranoside, followed by chromatography on DEAE-Sepharose, yielding 150 mg of protein/150 g wet weight of cells. Optical e.p.r. and low-temperature m.c.d. (magnetic circular dichroism) spectroscopies were used to investigate the nature of the protein ligands to the two haems in cytochrome bo from E. coli. Low-spin ferric haem b, the origin of a rhombic e.p.r. spectrum with g = 2.98, 2.26 and 1.50, gives rise to a charge-transfer band in the near-i.r. m.c.d. spectrum at 1622 nm. It is therefore concluded that haem b is co-ordinated by two histidine residues. The low-temperature m.c.d. spectrum of dithionite-reduced cytochrome bo comprises bands due both to low-spin ferrous haem b and to high-spin ferrous haem o. The bands arising from haem o show a direct correspondence with those in the m.c.d. spectrum of five-co-ordinate histidine-ligated ferrous haems such as myoglobin, implying that the protein residue liganding haem o is also histidine. This assignment was confirmed by measuring the e.p.r. spectrum of the nitric oxide derivative of fully reduced cytochrome bo. This showed a rhombic spectrum with g = 2.098, 2.008 and 1.987, and nuclear hyperfine splitting consistent with the co-ordination of ferrous haem by NO and histidine. The hyperfine splittings observed were 1.95 +/- 0.05 mT for the 14N of the NO ligand and 0.75 +/- 0.05 mT for the 14N of the proximal histidine. The e.p.r. spectrum of some samples of oxidized cytochrome bo show, at temperatures below 15 K, broad signals at g = 7.6, 3.6 and 2.8, and other preparations in the presence of glycerol yield signals at g = 10.8, 3.2 and 2.6. These signals, which are abolished by the addition of cyanide, are assigned to the binuclear centre, cytochrome o-CuB, suggesting that the binuclear site may display heterogeneity. Carbon monoxide reacts with the reduced enzyme with a stoichiometry of 1:1, and the dissociation constant for this reaction was determined to be 1.7 x 10(-6)M. The second-order rate constants for this reaction were measured and shown to be similar to those determined for bovine cytochrome aa3 [Gibson and Greenwood (1963) Biochem. J. 86, 541-554].

Dataset Information

Effects of inhibitory ligands on the aerobic carbon monoxide complex of cytochrome c oxidase.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets