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J B Chappell

Publications and source records attributed to J B Chappell.

At least 37 records · Page 2Linked to original sources
PubMed1975

The effects L-leucine on the synthesis of urea, glutamate and glutamine by isolated rat liver cells.

With either alanine or a mixture of 15 different amino acids as nitrogen source, the addition of L-leucine inhibited the synthesis of urea by isolated rat liver cells. With alanine present leucine promoted the production of glutamate and glutamine. Comparison of effects of leucine on soluble glutamate dehydrogenase, mitochondria and isolated cells supports the postulate that leucine exerts its effect through activation of glutamate dehydrogenase. It is suggested that this latter enzyme may not be as important for the production of NH3 for carbamoyl phosphate synthesis as has been considered hitherto.

Alanine↗
PubMed1974

The inhibition of mitochondrial dicarboxylate transport by inorganic phosphate, some phosphate esters and some phosphonate compounds.

1. P(i) competitively inhibited succinate oxidation by intact uncoupled mitochondria in the presence of sufficient N-ethylmaleimide to block the phosphate carrier, with a K(i) of 2.5mm. 2. Of a large number of phosphate esters and phosphonate compounds, phenyl phosphate and phenylphosphonate were found to inhibit competitively uncoupled succinate oxidation by intact but not broken mitochondria. By comparison, benzoate was a relatively weak competitive inhibitor of succinate oxidation by intact mitochondria but a relatively potent inhibitor of succinate dehydrogenase. 3. Phenyl phosphate and phenylphosphonate were non-penetrant, and inhibited P(i)-dependent swelling of mitochondria suspended in isosmolar ammonium malate in a manner non-competitive with P(i). The inhibitors did not affect mitochondrial swelling when tested with P(i) alone. 4. It is concluded that: (i) phenyl phosphate and phenylphosphonate behaved as non-penetrant analogues of P(i), since their inhibitory properties were in strict contrast with those of benzoate; (ii) phenyl phosphate and phenylphosphonate interacted with the dicarboxylate carrier but not with the phosphate carrier; (iii) P(i) was effective as a competitive inhibitor of succinate oxidation because of its being either an alternative substrate for the dicarboxylate carrier or competitive with succinate for the intramitochondrial cations as proposed by Harris & Manger (1968).

Adenosine Monophosphate↗
PubMed1974

The effects of acetylcolletotrichin on the mitochondrial respiratory chain.

1. Acetylcolletotrichin is a phytotoxic compound that has been isolated from the culture medium of the fungus Colletotrichum capsici (Grove et al., 1966). 2. With isolated liver and kidney mitochondria acetylcolletotrichin markedly inhibited the oxidation of succinate and those substrates with NAD-linked dehydrogenases, but did not inhibit the oxidation of ascorbate in the presence of tetramethyl-p-phenylenediamine. In this respect its action was similar to that of antimycin A. 3. Acetylcolletotrichin differed from antimycin in that, even at high concentrations which produced a maximal inhibitory effect, its action was partially reversed by uncoupling agents. Also acetylcolletotrichin had no detectable effect on the oxidative activity of blowfly flight-muscle mitochondria and was not very effective with heart mitochondria. 4. Acetylcolletotrichin inhibited the oxidative activity of liver mitochondria more markedly when respiration was stimulated by ADP together with phosphate and was less effective when respiration was stimulated by uncoupling agents. 5. There was an unusual interaction between the succinate oxidation system and the oxidation of glutamate together with malate. Thus, glutamate together with malate, even in the presence of rotenone, markedly decreased the effectiveness of acetylcolletotrichin in inhibiting succinate oxidation. 6. These effects were paralleled in the observed redox changes of cytochrome c. 7. The unusual behaviour of the cytochromes b in the presence of acetylcolletotrichin is described, and it is suggested tentatively that this inhibitor acts between cytochromes b with absorption maxima at 30 degrees C of approximately 560 and 565nm.

Adenosine Diphosphate↗
PubMed1974

Glutamate and aspartate transport in rat brain mitochondria.

1. Rat brain mitochondria did not swell in iso-osmotic solutions of ammonium or potassium (plus valinomycin) glutamate or aspartate, with or without addition of uncouplers. 2. Glutamate was able to reduce intramitochondrial NAD(P)(+); aspartate was able to cause partial re-oxidation. 3. These effects were inhibited by threo-hydroxy-aspartate in whole but not in lysed mitochondria. 4. The existence of a ;malate-aspartate shuttle' for the oxidation of extramitochondrial NADH was demonstrated. This shuttle requires the net exchange of glutamate for aspartate across the mitochondrial membrane. 5. Extramitochondrial glutamate did not inhibit intramitochondrial glutaminase under conditions in which the inhibition in lysed mitochondria was virtually complete. 6. The glutaminase activity of these mitochondria was not energy-dependent. 7. We conclude that these mitochondria do not possess a glutamate-hydroxyl antiporter similar to that of liver mitochondria nor a glutamate-glutamine antiporter similar to that of pig kidney mitochondria, but that they do possess a glutamate-aspartate antiporter.

Animals↗
PubMed1974

Changes in intramitochondrial adenine nucleotides in blowfly flight-muscle mitochondria.

1. With freshly isolated blowfly mitochondria 38% of the intramitochondrial adenine nucleotide was present as AMP. 2. On incubation with oxidizable substrates the AMP and ADP concentrations fell and that of ATP rose; with pyruvate together with proline the ATP concentration reached its maximum value at 6min; with glycerol phosphate the phosphorylation of endogenous nucleotide was more rapid. 3. Addition of the uncoupling agent carbonyl cyanide phenylhydrazone caused a rapid fall of ATP and a parallel rise in ADP, then ADP was converted into AMP. 4. This was in contrast with rat liver mitochondria endogenous AMP concentrations, which were always lower than those of blowfly mitochondria and changed little under different metabolic conditions. 5. Evidence is presented that adenylate kinase (EC 2.7.4.3) has a dual distribution in blowfly mitochondria, a part being located in the matrix space and a part in the space between the outer and inner mitochondrial membranes, as in liver and other mitochondria. 6. The possible regulatory role of changing AMP concentrations in the mitochondrial matrix was investigated. Partially purified pyruvate carboxylase (EC 6.4.1.1) and citrate synthase (EC 4.1.3.7) were inhibited 30% by 2mm-AMP, whereas pyruvate dehydrogenase (EC 1.2.4.1) was unaffected. 7. AMP activated the NAD(+)-linked isocitrate dehydrogenase (EC 1.1.1.41) activity of blowfly mitochondria in the absence of ADP, but in the presence of ADP, AMP caused inhibition. 8. It is suggested that AMP may exert a controlling effect on the oxidative activity of blowfly mitochondria.

Adenine Nucleotides↗
PubMed1974

Adaptive changes in the capacity of systems used for the synthesis of citrulline in rat liver mitochondria in response to high- and-low-protein diets.

1. Citrulline synthesis was measured in mitochondria from rats fed on a standard diet, a high-protein diet, or on glucose. 2. With NH(4)Cl as the nitrogen source the rate of citrulline synthesis was higher in mitochondria from rats fed on a high-protein diet than in those from rats fed on a standard diet. When rats were fed solely on glucose the rate of synthesis of citrulline from NH(4)Cl was very low. 3. With glutamate as the nitrogen source the relative rates of citrulline synthesis were much lower than when NH(4)Cl was present, but similar adaptive changes occurred. 4. The activity of the mitochondrial glutamate-transporting system increased two to three times on feeding rats on a high-protein diet, but the K(m) for glutamate was unchanged. 5. Adaptive changes in certain intramitochondrial enzymes were also measured. 6. The results were interpreted to indicate that when an excess of substrate was present, citrulline synthesis from NH(4)Cl was rate-limited by the intramitochondrial concentration of N-acetyl-glutamate, but citrulline synthesis from glutamate was rate-limited primarily by the activity of the glutamate-transporting system.

Ammonium Chloride↗
PubMed1973

The intramitochondrial location of the glutaminase isoenzymes of pig kidney.

1. The glutaminase activity of pig kidney is located almost entirely in the cortex. 2. Pig renal cortex contains two glutaminases, one phosphate-dependent and one phosphate-independent. Both isoenzymes are localized exclusively in the mitochondria. 3. After sonication of the mitochondria, the phosphate-dependent isoenzyme is entirely soluble, whereas approximately half the phosphate-independent isoenzyme is associated with the membranes. 4. In intact mitochondria, the activities of both isoenzymes respond to changes in the pH of the intramitochondrial compartment. 5. It is concluded that both glutaminase isoenzymes are situated in the intramitochondrial compartment, and that the phosphate-independent glutaminase may be bound to the inside of the inner mitochondrial membrane.

Animals↗
PubMed1973

Transport of glutamine and glutamate in kidney mitochondria in relation to glutamine deamidation.

1. In the absence of added ADP glutamine is transformed by pig kidney mitochondria to ammonium glutamate, which appears in the external medium. This reaction is stimulated only slightly by the addition of ADP, but under these conditions about 20% of the glutamate is oxidized to aspartate. 2. Externally added glutamate is oxidized to aspartate, and at about the same rate as glutamine. 3. The net rates of glutamine and glutamate influx into the intramitochondrial compartment are very slow. 4. The phosphate-dependent glutaminase activity of intact mitochondria is stimulated by the provision of energy. 5. The provision of energy also decreases the concentration of glutamate and increases the concentration of glutamine in the intramitochondrial compartment. These energy-linked changes in the glutamine and glutamate concentrations are of equal magnitude. 6. It is suggested that transport of glutamine and glutamate across the inner membrane of kidney mitochondria occurs by an obligatory exchange between the two metabolites, and is electrogenic. The existence of an electrogenic glutamine-glutamate anti-porter is proposed.

Adenosine Diphosphate↗
PubMed1973

Effect of L-leucine on the nitrogen metabolism of isolated rat liver mitochondria.

1. l-Leucine strongly activated intramitochondrial glutamate dehydrogenase in the direction of glutamate synthesis. 2. In the deamination direction, the enzyme was not stimulated by leucine. This was probably due to a rate-limiting transport of glutamate across the mitochondrial membrane. 3. The effect of leucine on the kinetic constants of glutamate dehydrogenase in a mitochondrial sonicate was studied. 4. In isolated mitochondria, leucine did not stimulate the synthesis of citrulline with glutamate as the source of NH(3). 5. Leucine very markedly stimulated the synthesis of glutamate from added 2-oxoglutarate+NH(4)Cl. 6. Under conditions where glutamate and citrulline could be synthesized simultaneously from added NH(4)Cl, leucine greatly increased glutamate synthesis at the expense of citrulline synthesis. 7. It is suggested that the intramitochondrial leucine concentration may be a factor influencing the nitrogen metabolism of the liver cell.

Ammonia↗
PubMed1973

Fluorocitrate inhibition of aconitate hydratase and the tricarboxylate carrier of rat liver mitochondria.

1. The effect of biologically synthesized and purified fluorocitrate on the metabolism of tricarboxylate anions by isolated rat liver mitochondria was investigated, in relation to the claim by Eanes et al. (1972) that this fluoro compound inhibits the tricarboxylate carrier at concentrations at which it has little effect on the aconitate hydratase activity. 2. That the inhibitory action of fluorocitrate is at the level of the aconitate hydratase and not at the level of the tricarboxylate carrier is indicated by the following findings. Although the oxidation of citrate and cis-aconitate, but not that of isocitrate, was inhibited by fluorocitrate, the exchange of internal citrate for external citrate or l-malate was not. Had the tricarboxylate carrier been affected, these latter exchange reactions would have been inhibited. 3. By using aconitate hydratase solubilized from mitochondria it was found that with citrate as substrate the inhibition by fluorocitrate was partially competitive (K(i)=3.4x10(-8)m), whereas with cis-aconitate as substrate the inhibition was partially non-competitive (K(i)=3.0x10(-8)m).

Aconitate Hydratase↗
PubMed1973

The transport of inorganic phosphate by the mitochondrial dicarboxylate carrier.

1. N-Ethylmaleimide inhibited the influx and efflux of P(i) in rat liver mitochondria. 2. The efflux was stimulated by either succinate or malate in the presence of N-ethylmaleimide, and this stimulation was reversed by 2-n-butylmalonate. 2-Oxoglutarate and citrate, even in the presence of low concentrations of malate, were relatively ineffective in stimulating efflux of P(i) under these conditions, as was glutamate. 3. By using radioactively labelled P(i) and dicarboxylate ions an exchange was demonstrated, the stoicheiometry of which was 1.3+/-0.5 dicarboxylate ions:1 P(i) (n=10). 4. An exchange between unlabelled and labelled P(i) in the presence of N-ethylmaleimide was found which was sensitive to 2-n-butylmalonate. 5. It is concluded that the mitochondrial dicarboxylate carrier can transport phosphate by an exchange diffusion with certain penetrant dicarboxylic acids or with phosphate itself. The exchange mechanism is sensitive to 2-n-butylmalonate but is unaffected by N-ethylmaleimide; the action of mersalyl in this context is commented on.

Animals↗
PubMed1973

Biogenesis of mitochondria. The effects of altered membrane lipid composition on cation transport by mitochondria of Saccharomyces cerevisiae.

1. The fatty acid composition of the membrane lipids of a fatty acid desaturase mutant of Saccharomyces cerevisiae was manipulated by growing the organism in a medium containing defined fatty acid supplements. 2. Mitochondria were obtained whose fatty acids contain between 20% and 80% unsaturated fatty acids. 3. Mitochondria with high proportions of unsaturated fatty acids in their lipids have coupled oxidative phosphorylation with normal P/O ratios, accumulate K(+) ions in the presence of valinomycin and an energy source, and eject protons in an energy-dependent fashion. 4. If the unsaturated fatty acid content of the mitochondrial fatty acids is lowered to 20%, the mitochondria simultaneously lose active cation transport and the ability to couple phosphorylation to respiration. 5. The loss of energy-linked reactions is accompanied by an increased passive permeability of the mitochondria to protons. 6. Free fatty acids uncouple oxidative phosphorylation in yeast mitochondria and the effect is reversed by bovine serum albumin. 7. The free fatty acid contents of yeast mitochondria are unaffected by depletion of unsaturated fatty acids, and free fatty acids are not responsible for the uncoupling of oxidative phosphorylation in organelles depleted in unsaturated fatty acids. 8. It is suggested that the loss of energy-linked reactions in yeast mitochondria that are depleted in unsaturated fatty acids is a consequence of the increased passive permeability to protons, and is caused by a change in the physical properties of the lipid phase of the inner mitochondrial membrane.

Biological Transport↗