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Biomedical subjects

F M Harold

Publications and source records attributed to F M Harold.

At least 55 records · Page 3Linked to original sources
PubMed1975

Accumulation of arsenate, phosphate, and aspartate by Sreptococcus faecalis.

Uptake of arsenate and phosphate by Streptococcus faecalis 9790 is strictly dependent on concurrent energy metabolism and essentially unidirectional. targinine supports uptake only in presence of glycerol or related substances; glycerol is not directly involved in transport but depletes the cellular orthophosphate pool and thus relieves feedback inhibition of transport. Uptake of phosphate and arsenate is stimulated by K+ and by other permeant cations. The results suggest that electroneutrality is preserved by compensatory movement of either H+ or OH minus. Ionophores and N,N'-dicyclohexylcarbodiimide, which prevent establishment of a proton motive force, block the accumulation of thiomethylgalactoside and of threonine but not that of arsenate or phosphate. We conclude that arsenate accumulation requires adenosine 5'-triphosphate but is not driven by the proton-motive force. However, conditions and reagents that lower the cytoplasmic pH do inhibit accumulation of arsenate and phosphate, suggesting that uptake depends on the capacity of the cells to maintain a neutral or alkaline cytoplasm. We therefore propose that phosphate accumulation is an electroneutral exchange for OH driven by adenosine 5'-triphosphate or by a metabolite thereof. Accumulation of aspartate and glutamate also requires adenosine 5'-triphosphate but not the proton-motive force and may involve a similar mechanism.

Adenosine Triphosphate↗
PubMed1974

Lactic acid translocation: terminal step in glycolysis by Streptococcus faecalis.

Streptococcus faecalis obtains metabolic energy chiefly from the conversion of glucose to lactic acid; the present experiments deal with the mechanism of lactic acid translocation across the cytoplasmic membrane. Efflux of [(14)C]lactate from preloaded cells was accelerated by raising the external pH, and also by the ionophores nigericin and valinomycin. These results suggest that lactate leaves the cell by an electroneutral process, presumably as lactic acid. Further evidence was obtained by studying the entry of [(14)C]lactate into nonmetabolizing cells. It appears that the membrane is essentially impermeable to the lactate anion, but allows passage of lactic acid. The most persuasive evidence is that, upon establishment of a pH gradient such that the cytoplasm was alkaline, l-[(14)C]lactate accumulated in the cells against the concentration gradient. Accumulation was transient, and dissipated in parallel with the collapse of the pH gradient. The concentration gradient attained at the peak was a function of the pH difference. Ionophores which are known to collapse a pH gradient, such as nigericin and valinomycin, abolished accumulation of l-lactate. We infer that lactic acid translocation, whether into the cells or outward, is an electroneutral process and for that reason the distribution of lactic acid across the membrane is a function of the pH of cytoplasm and medium. The specificity of translocation and its kinetic parameters suggest that it is mediated by a carrier of low specificity.

Carbon Radioisotopes↗
PubMed1973

Role of an electrical potential in the coupling of metabolic energy to active transport by membrane vesicles of Escherichia coli.

Membrane vesicles from E. coli can oxidize D-lactate and other substrates and couple respiration to the active transport of sugars and amino acids. The present experiments bear on the nature of the link between respiration and transport. Respiring vesicles were found to accumulate dibenzyldimethylammonium ion, a synthetic lipid-soluble cation that serves as an indicator of an electrical potential. The results suggest that oxidation of D-lactate generates a membrane potential, vesicle interior negative, of the order of -100 mV. In vesicles lacking substrate, an electrical potential was created by induction of electrogenic efflux of K(+) with the aid of the K(+) ionophores, valinomycin and monactin. These conditions induced transient accumulation by the vesicles of [(14)C]proline and other metabolites. Experiments with inhibitors and ionophores indicate that neither ATP nor the respiratory chain is involved; the electrical potential generated by K(+) efflux is coupled directly to the transport systems. The results verify two predictions derived from Mitchell's chemiosmotic hypothesis: respiring vesicles generate an electrical potential of the proper polarity and magnitude; and a membrane potential is in itself sufficient to drive the active transport of amino acids and other metabolites.

Ammonium Chloride↗
PubMed1970

Extrusion of sodium and hydrogen ions as the primary process in potassium ion accumulation by Streptococcus faecalis.

Glycolyzing cells of Streptococcus faecalis accumulate K(+) with concurrent extrusion of equivalent amounts of H(+) and Na(+). An attempt was made to clarify the retionship between the movements of Na(+) and K(+). Sodium was displaced from cells glycolyzing in the presence of ammonia, diethylamine, tris(hydroxymethyl)aminomethane, and other nitrogenous cations; by contrast, K(+) was completely retained. Accumulation of K(+) by heterologous exchange for Na(+) was not inhibited by antibiotics which facilitate diffusion of K(+) across the membrane, but was blocked by proton conductors. The results indicate that extrusion of Na(+) and H(+) from the cells is a primary, energy-linked process which generates an electrical potential (interior negative); K(+) accumulation occurs in response to this potential. Two mutants deficient in K(+) accumulation and retention were examined in terms of this model. One mutant is apparently defective in exchange of K(+) for H(+). In the other mutant, exchange of K(+) for Na(+) is impaired.

Amines↗
PubMed1969

Energy coupling in the transport of beta-galactosides by Escherichia coli: effect of proton conductors.

Escherichia coli accumulates thiomethyl-beta-d-galactoside against a concentration gradient under anaerobic conditions. The accumulation was abolished by carbonylcyanide m-chlorophenylhydrazone, tetrachlorosalicylanilide, 2,4 dinitrophenol, and other uncouplers of oxidative phosphorylation even though oxidative phosphorylation would not be expected to occur anaerobically. In the presence of the uncouplers, the beta-galactoside carrier remained functional and catalyzed equilibration of thiomethylgalactoside across the membrane. The uncouplers did not inhibit the generation of adenosine triphosphate or protein turnover, or the accumulation of alpha-methylglucoside and glycerol by phosphorylation. We conclude that, at least anaerobically, uncouplers of oxidative phosphorylation do not interfere with energy metabolism in general, but prevent the utilization of metabolic energy for the active transport of galactosides. The uncouplers also facilitate passage of protons across the membrane. Various hypotheses are considered to explain why a proton-impermeable membrane may be required for active transport of galactosides and other substrates.

Adenosine Triphosphate↗
PubMed1968

Effects of nigericin and monactin on cation permeability of Streptococcus faecalis and metabolic capacities of potassium-depleted cells.

At a concentration of 10(-6)m, nigericin and monactin inhibited growth of Streptococcus faecalis, and the inhibition was reversed by addition of excess K(+). In the presence of certain antibiotics, the cells exhibited increased permeability to certain cations; internal Rb(+) was rapidly lost by exchange with external H(+), K(+) Rb(+), and, more slowly, with Na(+) and Li(+). No effect was observed on the penetration of other small molecules. Cation exchanges induced by nigericin and monactin were metabolically passive and apparently did not involve the energy-dependent K(+) pump. When the cells were washed, the cytoplasmic membrane recovered its original impermeability to cations. By use of monactin, we prepared cells whose K(+) content had been completely replaced by other cations, and the metabolic characteristics of K(+)-depleted cells were studied. Cells containing only Na(+) glycolyzed almost as well as did normal ones and, under proper conditions, could accumulate amino acids and orthophosphate. These cells also incorporated (14)C-uracil into ribonucleic acid but incorporation of (14)C-leucine into protein was strictly dependent upon the addition of K(+). When K(+) or Rb(+) was added to sodium-loaded cells undergoing glycolysis, these ions were accumulated by stoichiometric exchange for Na(+). From concurrent measurements of the rate of glycolysis, it was calculated that one mole-pair of cations was exchanged for each mole of adenosine triphosphate produced.

Adenosine Triphosphate↗