Biomedical subjects
Publications and source records attributed to R Douce.
Spinach leaf mitochondria (C3 plant) were capable of oxidizing Glycine. This oxidation was linked to the mitochondrial electron transport chain, was coupled to three phosphorylation sites and was sensitive to electron transport inhibitors. In marked contrast however, neither mitochondria from dark grown plants nor mitochondria from C4 leaf plants were capable of oxidizing this amino acid.
The enzymatic synthesis of lysophosphatidic acid, phosphatidic acid, monoacylglycerol and diacylglycerol from sn-[14C]glycerol 3-phosphate occurs in purified chloroplasts. The results indicate that: (1) the chloroplast extract contains a soluble acylase (acyl-CoA: sn-glycerol 3-phosphate acyltransferase); (2) the envelope fraction contains an acyl-CoA synthetase, a bound acylase (acyl-CoA: acyl-sn glycerol 3-phosphate acyltransferase) and a phosphatidic acid phosphatase; without chloroplast extract in the incubation medium, the envelope is unable to incorporate sn-glycerol 3-phosphate into phosphatidic acid and diacylglycerol; addition of chloroplast extract to the incubation medium induced a fast increase of the incorporation of sn-glycerol 3-phosphate into phosphatidic acid and diacylglycerol; thylakoids being unable to incorporate sn-glycerol 3-phosphate (in presence or absence of soluble chloroplast extract in the incubation medium) our results indicate that the envelope of spinach chloroplast is the site of phosphatidic acid and diacylglycerol synthesis; (3) diacylglycerol actively synthesized by the envelope is also the substrate for the first galactosylation enzyme.
Yeast cationic ferricytochrome c was able to bind to the spinach (Spinacia oleracea) chloroplast envelope with a low affinity (Kd = 1.1 mum). The total amount of low affinity binding sites was of the order of 50 nmol cytochrome c mg(-1) protein. We gave the evidence that binding of ferricytochrome c to the envelope was electrostatic and that the envelope membranes were strongly negatively charged. Addition of yeast ferricytochrome c to a preparation of intact washed chloroplasts (class I) induced a strong agglutination of chloroplasts.
A procedure was described for preparing intact mitochondria from spinach (Spinacia oleracea L.) leaves. These mitochondria oxidized succinate, malate, pyruvate, alpha-ketoglutarate, and NADH with good respiratory control and ADP/O ratios comparable to those observed with mitochondria from other plant tissues. Glycine was oxidized by the preparations. This oxidation linked to the mitochondrial electron transport chain, was coupled to three phosphorylation sites and was sensitive to electron transport and phosphorylation inhibitors.Cyanide completely inhibited the oxidation of NADH. The oxidation of succinate, malate, and glycine was only partially inhibited.
The effects of copper deficiency on cell culture growth, cell respiration, mitochondrial oxidative properties, and electron transport chain have been studied with suspension-cultured sycamore cells (Acer pseudoplatanus L.). Within the range of the copper concentration studied (0.1-25 mug/1 of culture medium), the mean rate of cell division is independent of copper concentration. An initial copper concentration lower than 2 mug/1 limited the maximum density of population reached at the stationary phase of growth.On a protein basis, the uncoupled O(2) uptake rates were about the same for normal and copper-deficient cells. In contrast, the half-maximal inhibition of O(2) uptake rate was obtained at greater KCN concentration in the normal cells (20 muM) compared to copper-deficient cells (2 muM). Similar results were obtained with the normal and copper-deficient sycamore cell mitochondria.In the copper-deficient mitochondria, the concentration of the cytochrome aa(3) was less than 0.02 nmol/mg mitochondrial protein or 1/20 of the normal rate. The b- and c-type cytochrome content was invariant with copper depletion. It appeared that cytochrome aa(3) is present in large excess in normal cells. This work also indicated that cytochrome c is a very mobile molecule.
1. The inhibition of the ADP-stimulated respiration of potato mitochondria by carboxyatractyloside is relieved by high concentration of ADP or by the uncoupler carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP). Atractyloside is a much less potent inhibitor than carboxyatractyloside. The inhibition of the ADP-stimulated respiration required about 60-times more atractyloside than carboxyatractyloside. 2. [35S]carboxyatractyloside and [3H]bongkrekic acid bind to potato mitochondria with high affinity (Kd = 10 to 20 nM, n=0.6-0.7 nmol per mg protein). Added ADP competes with carboxyatractyloside for binding; on the contrary ADP increases the amount of bound bongkrekic acid. [3H]atractyloside binds to potato mitochondria with a much lower affinity (Kd=0.45 muM) than carboxyatractyloside or bongkrekic acid. 3. Bound [3H]atractyloside is displaced by ADP, carboxyatractyloside and bongkrekic acid. The displacement of bound [35S]carboxyatractyloside by bongkrekic acid and of bound [3H]bongkrekic acid by carboxyatractyloside is markedly increased by ADP. 4. Bongkrekic acid competes with [35S]carboxyatractyloside for binding. Addition of a small concentration of ADP considerably enhances the inhibitory effect of bongkrekic acid on [35S]carboxyatractyloside binding. 5. The adenine nucleotide content of potato mitochondria is of the order of 1 nmol per mg protein. ADP transport in potato mitochondria is inhibited by atractyloside 30- to 40-times less efficiently than by carboxyatractyloside.
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The effects of cyanide and 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) on photosynthesis and respiration of intact chlorophyllic moss (Funaria hygrometrica) spore was investigated. Thirty micromolar cyanide strongly inhibited dark respiration, was without effect on photosynthesis at high light intensities (above the saturation plateau values), and stimulated photosynthesis at low light intensities (below the saturation plateau values). Three hundred nanomolar DCMU inhibited the photosynthesis and was without effect, even under light conditions, on the dark respiration. It seems likely, therefore, that in the chlorophyllic moss spore the cytochrome oxidase pathway is not functioning under high light intensities unless the photosynthesis is inhibited by DCMU.
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The envelope of the spinach chloroplast is a yellow membrane system with a unique carotenoid composition. Envelopes prepared from dark-treated leaves had a violaxanthin content up to 3.5 times the lutein plus zeaxanthin content, whereas in chloroplast envelopes from illuminated leaves this ratio was only 0.75. Light-catalyzed changes in violaxanthin content also occurred in the thylakoid fraction.The role of this reversible light-catalyzed de-epoxidation of violaxanthin in the function of the envelope of the chloroplast is discussed.
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