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E Hurt

Publications and source records attributed to E Hurt.

64 records · Page 4Linked to original sources
PubMed1983

Partial structure of the active moiety of a lipoprotein complexing proteoglycan from human aorta.

Proteoglycans and glycosaminoglycans of the intima-media extracellular matrix have been stated to play a role in lipoprotein deposition associated with atherogenesis. It is therefore important to characterize the active lipoprotein-complexing moiety of these macromolecular aggregates. We have isolated a soluble proteoglycan aggregate of approximately 5 X 10(6) molecular weight after homogenization of human aortic intima-media in an isosmotic sucrose solution, sequential differential centrifugation, dialysis, exclusion chromatography and preparative electrophoresis. This proteoglycan aggregate, labelled lipoprotein-complexing proteoglycan (LCP), has been previously shown to form specific complexes with low density lipoproteins, either isolated or in sera. Density gradient centrifugation in dissociative conditions of the LCP, cellulose acetate acetate electrophoresis of the subfractions, chondroitinases treatment and high performance liquid chromatography of the unsaturated disaccharides indicated that the glycosaminoglycan moiety was composed of 56% chondroitin-6-SO4, 26% hyaluronate and/or undersulfated chondroitin and 17% chondroitin-4-SO4. In pore-gradient polyacrylamide gel electrophoresis, the hyaluronate monomer appeared to have a molecular weight of 250000 while that of the chondroitin sulfates ranged between 50000 and 70000 after extensive treatment with protease. The fractions enriched in the chondroitin sulfate monomers were the most reactive towards LDL and their reactivity was abolished by chondroitinase AC indicating that the lipoprotein-complexing capacity of the LCP aggregate is associated to these molecules.

Aorta↗
PubMed1982

Reduction of cytochromes b6 and f in isolated plastoquinol-plastocyanin oxidoreductase driven by photochemical reaction centers from Rhodopseudomonas sphaeroides.

Photochemical reaction centers isolated from the bacterium Rhodopseudomonas sphaeroides are able to donate electrons to cytochromes b6 and f in the plastoquinol,-platocyanin oxidoreductase isolated from spinach chloroplasts. The reduction reactions occur only after the second single turnover flash, in a reaction which is sensitive to inhibitors of the reactions in the chloroplast membranes. When all the components of the b6f complex are oxidized prior to activation, both cytochromes b6 and f are reduced after the second flash. If cytochrome f is reduced prior to activation, cytochrome b6 is still reduced after the second flash, but as the potential is lowered so that the Rieske iron-sulfur cluster is reduced prior to activation, the reduction of cytochrome b6 fails. The b6f complex thus functions in such a way that a two-electron redox couple, probably a quinone, is capable of reducing both cytochrome b6 and cytochrome f, the latter via the Rieske iron-sulfur cluster, in a coupled reaction where both electrons must leave the two-electron carrier. Cytochrome b6 is thus reduced in a manner analogous to "oxidant-induced reduction."

Bacterial Proteins↗
PubMed1982

Identification of the polypeptides in the cytochrome b6/f complex from spinach chloroplasts with redox-center-carrying subunits.

An improved procedure for the isolation of the cytochrome b6/f complex from spinach chloroplasts is reported. With this preparation up to tenfold higher plastoquinol-plastocyanin oxidoreductase activities were observed. Like the complex obtained by our previous procedure, the complex prepared by the modified way consisted of five polypeptides with apparent molecular masses of 34, 33, 23, 20, and 17 kD, which we call Ia, Ib, II, III, and IV, respectively. In addition, one to three small components with molecular masses below 6 kD were now found to be present. These polypeptides can be extracted with acidic acetone. Cytochrome f, cytochrome b6, and the Rieske Fe-S protein could be purified from the isolated complex and were shown to be represented by subunits Ia + Ib, II, and III, respectively. The heterogeneity of cytochrome f is not understood at present. Estimations of the stoichiometry derived from relative staining intensities with Coomassie blue and amido black gave 1:1:1:1 for the subunits Ia + Ib/II/III/IV, which is interesting in of the presence of two cytochromes b6 per cytochrome f. Cytochrome f titrated as a single-electron acceptor with a pH-independent midpoint potential of +339 mV between pH 6.5 and 8.3, while cytochrome b6 was heterogeneous. With the assumption of two components present in equal amounts, two one-electron transitions with Em(1) = 40 mV and Em(2) = -172 at pH 6.5 were derived. Both midpoint potentials were pH-dependent.

Chloroplasts↗
PubMed1982

A cytochrome b/c1 complex with ubiquinol--cytochrome c2 oxidoreductase activity from Rhodopseudomonas sphaeroides GA.

A cytochrome b/c1 complex which catalyses the reduction of cytochrome c by ubiquinol has been isolated from Rhodopseudomonas sphaeroides GA. It contains two hemes b and substoichiometric amounts of ubiquinone-10 and of the Rieske Fe-S center per cytochrome c1, and is essentially free of reaction center and bacteriochlorophyll. The complex consists of three major polypeptides with apparent molecular masses of 40, 34 and 25 kDa. The 34-kDa polypeptide carries heme. Cytochrome c1 has a midpoint potential of 285 mV. For cytochrome b two midpoint potentials, at 50 and -60 mV, at pH 7.4, can be derived if one assumes two components of equal amount. Ubiquinol--cytochrome c oxidoreductase activity is specific for ubiquinol and bacterial cytochromes c, and is inhibited by antimycin A and 5-n-undecyl-6-hydroxy-4,7-dioxobenzothiazole. The complex shows oxidant-induced reduction of cytochrome b.

Chemical Phenomena↗
PubMed1981

A cytochrome f/b6 complex of five polypeptides with plastoquinol-plastocyanin-oxidoreductase activity from spinach chloroplasts.

The isolation of a cytochrome f/b6 complex from spinach chloroplasts, with high yield and purity is reported. The complex consists of five polypeptides with a molecular mass of 34, 33, 23.5, 20 and 17.5 kDa, and contains one cytochrome f, two cytochromes b6 and the Rieske Fe-S center with two non-heme irons. It does not contain plastocyanin and is almost completely devoid of chlorophyll and carotenoids, but lipid and detergent are present. It is lacking cytochrome b-559, although three of the five polypeptides seem to carry heme groups. The preparation has plastoquinol-plastocyanin oxidoreductase activity with plastoquinol-1 and plastoquinol-9, which is sensitive to 2,5-dibromomethylisopropyl-p-benzoquinone, to 2-iodo-6-isopropyl-3-methyl-2',4',4'-trinitrodiphenyl ether, to 5-n-undecyl-6-hydroxy-4,7-dioxobenzothiazole, and to bathophenanthroline. Characteristics of this activity with respect to substrate concentrations, pH, detergent effect and other parameters are described.

Chloroplasts↗
PubMed1980

Structural requirements of quinone coenzymes for endogenous and dye-mediated coupled electron transport in bacterial photosynthesis.

Electron transport in continuous light has been investigated in chromatophores of Rhodopseudomonas capsulata. Ala pho+, depleted in ubiquinone-10 and subsequently reconstituted with various ubiquinone homologs and analogs. In addition the restoration of electron transport in depleted chromatophores by the artificial redox compounds N-methylphenazonium methosulfate and N,N,N',N'-tetramethyl-p-phenylenediamine was studied. The following pattern of activities was obtained: (1) Reconstitution of cyclic photophosphorylation with ubiquinone-10 was saturated at about 40 ubiquinone molecules per reaction center. (2) Reconstitution by ubiquinone homologs was dependent on the length of the isoprenoid side chain and the amount of residual ubiquinone in the extracted chromatophores. If two or more molecules of ubiquinone-10 per reaction center were retained, all homologs with a side chain longer than two isoprene units were as active as ubiquinone-10 in reconstitution, and the double bonds in the side chain were not required. If less than two molecules per reaction center remained, an unsaturated side chain longer than five units was necessary for full activity. Plastoquinone, alpha-tocopherol, and naphthoquinones of the vitamin K series were relatively inactive in both cases. (3) All ubiquinone homologs, also ubiquinone-1 and -2, could be reduced equally well by the photosynthetic reaction center, as measured by light-induced proton binding in the presence of antimycin A and uncoupler. Plastoquinone was found to be a poor electron acceptor. (4) Photophosphorylation could be reconstituted by N-methylphenazonium methosulfate as well as by N,N,N',N'-tetramethyl-p-phenylenediamine in an antimycin-insensitive way, if more than two ubiquinones per reaction center remained. These compounds were active also in more extensively extracted particles reconstituted with ubiquinone-1, which itself was inactive.

Adenosine Triphosphate↗
PubMed1979

Vectorial redox reactions of physiological quinones. I. Requirement of a minimum length of the isoprenoid side chain.

Physiological quinones carrying isoprenoid side chains have been compared with homologues lacking the side chain, for their ability to carry electrons and protons from dithionite to ferricyanide, trapped in liposomes. Six differential observations were made: (1) Plastoquinone and ubiquinones, with a side chain of more than two isoprene units, are by far better mediators than their short-chain homologues. Also other benzoquinones lacking a long side chain are poor catalysts, except dimethyl-methylenedioxy-p-benzoquinone, a highly autooxidizable compound. Tocopherol is a good catalyst. (2) Vitamin K-1 and K-2 are poor mediators compared to vitamin K-3. (3) The reaction catalyzed by quinones carrying long isoprenoid side chains has an about three-fold higher activation energy, irrespective of the catalytic efficiency. (4) The reaction catalyzed by quinones lacking a long side chain follows pseudo first-order kinetics, while the reaction with quinones carrying a long side chain is of apparently higher order. (5) The rate with ubiquinone-1 is increasing pH, while with ubiquinone-9 it is decreasing. (6) The reaction mediated by short-chain quinones seems to be satuarated at lower dithionite concentration. We conclude that isoprenoid quinones are able to translocate electrons and protons in lipid membranes, and that the side chain has a strong impact on the mechanism. This and the relevance of the model reaction for electron and proton transport in photosynthesis and respiration is discussed.

Energy Transfer↗