PubMed Health⌕ Search

Biomedical subjects

R W Hendler

Publications and source records attributed to R W Hendler.

66 records · Page 4Linked to original sources
PubMed1972

Respiration and protein synthesis in Escherichia coli membrane-envelope fragments. IV. Chemical and cytological characterization and biosynthetic capabilities of fragments obtained by mild procedures.

Membrane-envelope fragments have been isolated from Escherichia coli by comparatively mild techniques. The use of DNAase, RNAase, detergents, sonication, lysozyme, and ethylenediaminetetraacetate were avoided in the belief that rather delicate, but metabolically important, associations may exist between the plasma membrane and various cytoplasmic components. The membrane-envelope fragments have been characterized in terms of their content of major chemical components as well as their electron microscope appearance. Fractions containing membrane-envelope fragments were found to possess appreciable DNA- and protein-synthesizing activities. The fragments were rich in membrane content as determined by reduced nicotinamide adenine dinucleotide (NADH) oxidase activity and deficient in soluble components as measured by NADH dehydrogenase activity. The particulate fraction obtained between 20,000 g and 105,000 g and usually considered a ribosomal fraction was rich in membrane content and had a relatively high capacity for DNA synthesis. Envelope fragments sedimenting at 20,000 g attained very high levels of incorporation of amino acids into protein.

Amino Acids↗
PubMed1972

Respiration and protein synthesis in Escherichia coli membrane-envelope fragments. VI. Solubilization and characterization of the electron transport chain.

Membranes obtained from Escherichia coli have been solubilized with deoxycholate. The solubilized dehydrogenases and cytochromes are not sedimented at 105,000 g. These components readily penetrate the "included space" of Sepharose 4B (Pharmacia Fine Chemicals Inc., Uppsala, Sweden) and polyacrylamide gels and have been fractionated on the basis of molecular size. Solubilization destroys nicotinamide adenine dinucleotide, reduced form (NADH) oxidase and D-lactate oxidase activities, but leaves an appreciable part of the original succinoxidase activity intact. Evidence for a succinate dehydrogenase-cytochrome b(1) complex is given. Menadione added to the solubilized preparation does not elicit NADH oxidase activity nor stimulate the existing succinoxidase activity, but does provoke an active D-lactate oxidase activity. This D-lactate oxidase activity, however, does not use cytochromes and is not sensitive to cyanide.

Bacterial Proteins↗
PubMed1971

Respiration and protein synthesis in Escherichia coli membrane-envelope fragments. V. On the reduction of nonheme iron and the cytochromes by nicotinamide adenine dinucleotide and succinate.

The sensitivity of nicotinamide adenine dinucleotide (NADH) oxidase and succinoxidase to metal chelators, the generation of an electron paramagnetic resonance (EPR) signal upon addition of these substrates, and the rate of formation of the EPR signal relative to the rate of the cytochrome reduction suggest the participation of nonheme iron proteins in the respiratory process of Escherichia coli. The most inhibitory metal chelator, thenoyltrifluoro acetone, inhibited the reduction of nonheme iron and cytochromes but did not prevent the reoxidation of the reduced forms. The EPR signal, dehydrogenase, and oxidase activities evoked by NADH are considerably greater than the corresponding activities evoked by succinate. Because both substrates can reduce almost all of the cytochromes, a model in which fewer succinate dehydrogenase-nonheme iron protein complexes are linked to a common cytochrome chain than NADH dehydrogenase-nonheme iron protein complexes is considered likely.

Acetone↗
PubMed1970

Respiration and protein synthesis in Escherichia coli membrane-envelope fragments. II. Effects of fatty acids and albumin on respiration.

Fatty acids inhibited the ability of Escherichia coli membrane-envelope fragments to catalyze the oxidation of succinate and nicotinamide adenine dinucleotide, reduced form (NADH) and also inhibited the response of the Clark oxygen electrode to nonenzymatic oxygen uptake. In all cases, unsaturated fatty acids were much more inhibitory than saturated fatty acids. Albumin afforded complete protection from inhibition in the nonenzymatic oxygen-uptake experiments but only partial protection for the respiratory activities of the membrane fragments. The succinoxidase activity was totally inhibited by bovine serum albumin at concentrations that inhibited succinate dehydrogenase only slightly and NADH oxidase not at all. The E. coli acellular preparation showed no dehydrogenase or oxidase activity for any of the fatty acids under a variety of conditions. These conditions included variations of pH, concentration of fatty acids, and the presence or absence of albumin, CoA, ATP, NAD, cysteine, succinate, and carnitine. It thus appears that E. coli grown in the absence of fatty acid can not use fatty acids as an energy source.

Adenosine Triphosphate↗
PubMed1970

Respiration and protein synthesis in Escherichia coli membrane-envelope fragments. 3. Electron microscopy and analysis of the cytochromes.

The membranous nature of pellets obtained from broken Escherichia coli spheroplasts by successive centrifugation at 3500 g (P(1)), 20,000 g (P(2)), and 105,000 g (P(3)), has been established by electron microscopy. Spectrophotometric analysis has shown that about 90% of the cytochromes are concentrated in the particulate fractions. The crude ribosomal pellet (P(3)) contained as much of the total cytochromes as did the pellet obtained at 20,000 g (P(2)). The high cytochrome content of P(3) is consistent with its high oxidative activity (1) and the presence of membrane vesicles in this fraction. Analysis at 77 degrees K intensified the optical extinction of all the cytochrome absorption bands, but the degree of intensification was not uniform for each fraction nor for each band within a given fraction. Carbon monoxide had little or no inhibiting effect on NADH oxidation. Reduced plus carbon monoxide difference spectra yielded artifactual absorption bands in the wave length regions where reduced vs. oxidized absorption bands normally occur. Succinate and NADH, either together or separately, reduced nearly all of the cytochromes, indicating that the cytochrome portion of the electron-transport chain is shared by both substrates. A tentative formulation of the electron-transport chain is presented.

Carbon Monoxide↗
PubMed1969

Respiration and protein synthesis in Escherichia coli membrane-envelope fragments. I. Oxidative activities with soluble substrates.

This paper describes experiments conducted with membranous and soluble fractions obtained from Escherichia coli that had been grown on succinate, malate, or enriched glucose media. Oxidase and dehydrogenase activities were studied with the following substrates: nicotinamide adenine dinucleotide, reduced form (NADH), nicotinamide adenine dinucleotide phosphate, reduced form (NADPH), succinate, malate, isocitrate, glutamate, pyruvate, and alpha-ketoglutarate. Respiration was virtually insensitive to poisons that are commonly used to inhibit mitochondrial systems, namely, rotenone, antimycin, and azide. Succinate dehydrogenase and NADH, NADPH, and succinate oxidases were primarily membrane-bound whereas malate, isocitrate, and NADH dehydrogenases were predominantly soluble. It was observed that E. coli malate dehydrogenase could be assayed with the dye 2,6-dichlorophenol indophenol, but that porcine malate dehydrogenase activity could not be assayed, even in the presence of E. coli extracts. The characteristics of E. coli NADH dehydrogenase were shown to be markedly different from those of a mammalian enzyme. The enzyme activities for oxidation of Krebs cycle intermediates (malate, succinate, isocitrate) did not appear to be under coordinate genetic control.

Bacterial Proteins↗