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Biomedical subjects

M Wikström

Publications and source records attributed to M Wikström.

At least 199 records · Page 11Linked to original sources
PubMed1984

Activated polymorphonuclear leucocytes consume vitamin C.

Polymorphonuclear leucocytes (PMN) are known to produce superoxide and other oxygen derivatives upon activation as part of their microbicidal armory. Here we report that extracellular ascorbate is effectively oxidised by activated but not by resting human PMN in vitro. The oxidation of ascorbate is mainly caused by the superoxide that is generated by the activated cells, as shown by its effective inhibition by superoxide dismutase. However, myeloperoxidase, which may generate hypochlorite, also contributes to a significant extent. Ascorbate reduces superoxide to peroxide, as indicated by measurements of the stoichiometry of ascorbate and oxygen consumption. These results support the notion that extracellular ascorbate may serve as an important physiological protecting agent against oxygen radical damage in inflammation.

Ascorbic Acid↗
PubMed1984

Two protons are pumped from the mitochondrial matrix per electron transferred between NADH and ubiquinone.

The spectrophotometric indicators neutral red and safranine were used to determine the relative H+/e- ratios of proton uptake from the mitochondrial matrix, and the q+/e- ratios of electrical charge translocation, during oxidation of beta-OH-butyrate and succinate by ferricyanide in rat liver mitochondria. With beta-OH-butyrate both ratios were higher than with succinate by a factor close to 3.0. Since there is full agreement that H+/e- of proton uptake and q+/e- of charge translocation are both equal to unity for oxidation of succinate (or ubiquinol) by ferricytochrome c, the corresponding ratios for oxidation of NADH by ubiquinone and cytochrome c are near 2.0 and 3.0, respectively.

Animals↗
PubMed1984

Calorimetric studies of cytochrome oxidase-phospholipid interactions.

Thermotropic phase transitions in phospholipid vesicles reconstituted with mitochondrial cytochrome oxidase (EC 1.9.3.1) were studied using differential scanning calorimetry. Both dimyristoylphosphatidylcholine (DMPC) and mixtures of DMPC and cardiolipin were used at different lipid-to-protein ratios. The incorporated protein reduces the energy absorbed during phase transitions of DMPC vesicles, and causes a small decrease in the transition temperature (tm). delta H depends on the amount of protein in the vesicles. This dependence indicates that about 72 DMPC molecules are influenced per cytochrome alpha alpha 3 monomer. The transition parameters remain unaffected by changes in ionic strength or by reduction of the enzyme. Incorporation of cytochrome oxidase depleted of subunit III into DMPC liposomes resulted in a larger decrease of tm, but the amount of perturbed phospholipids remains similar to that in the case of the intact enzyme. Incorporation of cytochrome oxidase into DMPC/cardiolipin vesicles counteracts the effect of cardiolipin in decreasing the enthalpy of the DMPC transition. Thus cytochrome oxidase segregates the phospholipids by attracting cardiolipin from the bulk lipid. Cytochrome c does not significantly affect this apparent cardiolipin 'shell' around membranous cytochrome oxidase.

Animals↗
PubMed1983

Monoclonal antibodies to cholera toxin with special reference to cross-reactions with Escherichia coli heat-labile enterotoxin.

Seventy monoclonal antibodies to cholera toxin were prepared and characterized. All were of immunoglobulin G (IgG) isotypes (39 IgG1, 29 IgG2, and 2 IgG3). A total of 61 clones produced antibody directed against the B subunit, and 9 clones produced antibodies with specificity for the cholera toxin A subunit. Among both the anti-B and anti-A antibodies, there were representatives which showed full cross-reactivity with the heat-labile enterotoxin of Escherichia coli (14 clones), others which gave partial cross-reactions (12), and still others (44) which did not cross-react. Although 24 of 25 tested anti-B monoclonal antibodies could neutralize cholera toxin, none of the 9 anti-A clones had any detectable neutralizing ability. Among the anti-B antibodies, those which cross-reacted completely with E. coli heat-labile enterotoxin all had strong cholera toxin-neutralizing capacity, whereas those with lesser or no degree of cross-reactivity varied more in their neutralizing potency. The isolation of monoclonal antibodies that distinguish between enterotoxins of different bacterial origin suggests the possibility of developing immunodiagnostic methods allowing species-specific enterotoxin detection in stools of patients with diarrheal disease.

Antibodies, Monoclonal↗
PubMed1983

Production of histolytic enzymes by a combination of oral bacteria with known pathogenicity.

Eight oral bacterial strains, isolated from an infected root canal, have been investigated for their capacity to produce histolytic enzymes. The determination was performed using methods espoused by two different principles. Eleven out of 12 enzymes examined were demonstrated in the "eight-strain collection". In no single bacterial strain were all enzymes revealed. It was suggested that the pathogenicity of the bacterial strains, singly or in combination, was not solely dependent on the production of these enzymes. The histolytic enzymes may have a potentiating role on other pathogenic factors in infectious diseases.

Bacteria↗
PubMed1983

The present state of the chemiosmotic coupling theory.

Although the general principles of the chemiosmotic coupling theory have become widely accepted, the (degree of) loc(aliz)ation of electrochemical proton potential difference cannot yet be deduced from the existing experimental data. Many results are not in ready accordance with the idea that one protonic electrochemical potential difference, i.e. the one between a homogeneous inner and a homogeneous outer aqueous phase, would be the high-free-energy intermediate of membrane-linked free-energy transduction. Rather, free-energy transduction in an organelle like a mitochondrion or a chloroplast might take place in large number (about 1 per H+-ATPase) of miniature chemiosmotic systems. The energized protons produced in such a miniature system might be largely (but not totally) confined to a proton-domain belonging to it. Hence, there might be many (rather than one) different relevant proton gradients.

Chloroplasts↗
PubMed1981

Energy-dependent reversal of the cytochrome oxidase reaction.

Energization of isolated rat liver mitochondria with ATP under conditions in which cytochrome c is poised in a highly oxidized state shifts the state of cytochrome oxidase (cytochrome c oxidase; ferrocytochrome c:oxygen oxidoreductase, EC 1.9.3.1) from fully oxidized to two new spectroscopically distinguishable states depending on the applied phosphorylation potential and redox potential at cytochrome c. Both new states are spectrally similar or identical to two previously described intermediates in the reaction between reduced enzyme and O2. The data suggest that the energy-dependent transitions are due to reversed electron transfer from water to ferricytochrome c linked to accumulation of intermediates of O2 reduction at the catalytic heme a3/copper center. Titrations with redox potential indicate that each transition is a one-electron step, a finding that would identify the second observed compound as enzyme-bound peroxide or its equivalent. This is consistent with this compound being spectrally identical to "Compound C," previously described as the reaction product between half-reduced oxidase (two electrons) and O2. On the basis of these data a catalytic scheme of O2 reduction is proposed for the heme a3/copper center of cytochrome oxidase.

Adenosine Diphosphate↗
PubMed1981

Quaternary structure of bovine cytochrome oxidase.

A hydrodynamically homogeneous preparation of bovine mitochondrial cytochrome c oxidase can be obtained by anion-exchange chromatography of alkaline-treated enzyme, followed by a gel permeation chromatography step, which further removes some (aggregated) apoprotein. The molecular weight, Mr, of the monodisperse enzyme in Triton X-100 was found to be 210000. This complex is composed of six different polypeptides, with Mr summing up to about 110000 in toto, in a relative one-to-one stoichiometry. Two sets of these subunits constitute the 210000-Mr enzyme complex. In contrast to our earlier report [Saraste, Penttilä, Coggins, and Wikström, FEBS Lett. 114 (1980) 35-38] the 210000-Mr enzyme contains four (and not two) haems A, and therefore represents the dimer of cytochrome aa3. One of the proposed seven subunits, number III, is lacking in this enzyme preparation.

Animals↗
PubMed1980

Effect of 2-n-heptyl-4-hydroxyquinoline N-oxide on proton permeability of the mitochondrial membrane.

The respiratory-chain inhibitor 2-n-heptyl-4-hydroxyquinoline N-oxide catalyses transmembrane proton transport driven by a pH gradient in isolated rat liver mitochondria. This effect explains the apparent blockade of net proton translocation by this compound in mitochondria respiring with ferrocyanide as described by Papa, Lorusso, Guerrieri, Boffoli, Izzo & Capuano [(1977) in Bioenergetics of Membranes (Packer, Papageorgiu & Trebst, eds.), pp. 377-388, Elsevier/North-Holland, Amsterdam] and by Lorusso, Capuano, Boffoli, Stefanelli & Papa [(1979) Biochem. J. 182, 133-147].

Animals↗
PubMed1980

Interactions of Ca2+ and H+ with heme A in cytochrome oxidase.

Ca2+ ions shift the absorption spectrum of reduced cytochrome a in mitochondria by acting from the outside of the membrane. In isolated cytochrome oxidase the shift may be induced by either Ca2+ or H+, the apparent pK varying between 6.20 and 5.75 depending on the state of cytochrome a3. Studies of the Soret band show that Ca2+ also shifts the spectrum of ferrocytochrome a3 in isolated oxidase in contrast to the situation in mitochondria or isolated oxidase reconstituted into liposomes. Model studies with reduced bis-imidazole heme A reveals an analogous spectral shift induced by Ca2+. Esterification of the propionate carboxyls of heme A abolishes the spectral shift, suggesting that it is due to interaction of Ca2+ with these groups. When taken together with the data with intact mitochondria, this suggests that the propionate side chains of cytochrome a are accessible to Ca2+ and H+ from the outside of the mitochondrial membrane. In the soluble enzyme both hemes a and a3 are accessible. Thus heme a may be located near the outside of the inner membrane whereas heme a3 experiences a different environment in which no Ca2+ shift occurs.

Animals↗
PubMed1979

On the stoichiometry and thermodynamics of proton-pumping cytochrome c oxidase in mitochondria.

Different approaches have been used to evaluate the stoichiometry of proton translocation linked to cytochrome c oxidase in rat liver mitochondria. A mathematical model was designed that successfully describes the kinetics of redox-linked proton translocation provided that the rate of electron transfer is not too high. With ascorbate as reductant, an essentially pH-independent (in the pH range 6--8.5) proton ejection stoichiometry (H+/e-) is obtained from either initial rates of H+ ejection (0.86 +/- 0.12), or the model (0.87 +/- 0.14). Similar results are obtained with either ferrocyanide, N.N.N',N'-tetramethyl-p-phenylenediamine or externally added cytochrome c mediating between ascorbate and cytochrome c in rotenone- and antimycin-inhibited mitochondria. Oxygen pulse experiments with ferrocytochrome c as substrate show fully uncoupler-sensitive redox-linked proton ejection with a stoichiometry of 0.78 +/- 0.14. With murexide to measure Ca2+ uptake during oxidation of ferrocyanide, we found a stoichiometry of two positive charges taken up/electron transferred, confirming earlier findings. These results provide strong evidence that cytochrome c oxidase functions as a redox-linked proton pump with a stoichiometry of one H+ ejected and two charges translocated/electron transferred. The thermodynamic consequences of the proton pump are discussed and a maximal P/O ratio of 1 1/3 for 'site 3' is predicted in agreement with state 4 redox potentials and phosphate potential.

Animals↗