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J Swierczynski

Publications and source records attributed to J Swierczynski.

41 records · Page 3Linked to original sources
PubMed1976

Regulation of alpha-glycerophosphate dehydrogenase activity in human term placental mitochondria.

1. alpha-Glycerophosphate dehydrogenase (sn-glycerol-3-phosphate:(acceptor) oxidoreductase, EC 1.1.99.5) activity in mitochondria isolated from human term placenta was found to be inhibited by ethyleneglycolbis (beta-aminoethyl ether)-N,N'-tetraacetic acid (EGTA). Addition of an excess of calcium ions to the incubation medium completely restored the original activity. The concentration of free calcium ion required to activate the alpha-glycerophosphate dehydrogenase was found to vary between 10 and 100 nM. 2. The pH optimum for alpha-glycerophosphate dehydrogenase activity varied with substrate concentration. The pH optima were 7.4 and 8.0 in the presence of 2 or 8 mM alpha-glycerophosphate, respectively. The apparent Km for alpha-glycerophosphate also varied with pH; the values being 0.4 mM at pH 7.05, 1.5 mM at pH 7.8, and 3.5 mM at pH 8.5. 3. alpha-Glycerophosphate dehydrogenase activity was inhibited by palmitoyl-CoA in a competitive manner with an apparent Ki value of about 10 muM. This inhibition was less pronounced in the presence of calcium or magnesium ions. 4. The activity of alpha-glycerophosphate dehydrogenase was inhibited by phosphoenolpyruvate, D- and DL-glyceraldehyde 3-phosphate and 3-phosphoglyceric acid, in a competitive manner, the apparent Ki values being 0.5, 0.95, 0.12 and 1.5 mM, respectively. 5. alpha-Glycerophosphate dehydrogenase activity in human placental mitochondria was found to be more sensitive to phosphoenolpyruvate, than the activity of the same enzyme in rat skeletal muscle mitochondria. alpha-Glycerophosphate dehydrogenase activity in rat brown adipose tissue mitochondria was only slightly affected by phosphenolpyruvate under the same conditions. 6. The data obtained suggest that the activity of alpha-glycerophosphate dehydrogenase in human placental mitochondria may be controlled by changes of the cytosolic level of palmitoyl-CoA, some glycolytic intermediates, and pH.

Adipose Tissue, Brown↗
PubMedDate not supplied

Isolation, properties and role in progesterone biosynthesis of cytosolic malic enzyme from human term placenta.

Cytosolic malic enzyme (L-malate: NADP oxidoreductase decarboxylating, EC I. I. I.40) has been isolated and purified from postmitochondrial supernatant of human term placenta by ammonium sulphate fractionation, chromatography on diethylaminoethyl- (DEAE-)cellulose, Sepharose 6B, ADP-Sepharose 4B and Ultrogel AcA-34 to apparent homogeneity as judged from polyacrylamide gel electrophoresis (PAGE). The specific activity of the purified enzyme was 24.0 mumol X min-1 X mg-1 protein, which corresponds to about 7500-fold purification. The molecular weight of the native enzyme was determined by gel filtration to be about 250,000. Sodium dodecyl sulphate- (SDS-)PAGE showed one polypeptide band of molecular weight 63,000. It appears that the native protein is a tetramer composed of subunits of identical molecular weight. The isoelectric point of the purified malic enzyme was pH 5.55. The enzyme was shown to carboxylate pyruvate in the presence of high concentrations of pyruvate and bicarbonate at about 80 per cent of the rate of the forward reaction. The optimum pH for the carboxylation reaction was pH 7.3, and that for the decarboxylation reaction varied with malate concentration. The Km values, determined at pH 7.2, for malate, NADP+, Mn2+, and Mg2+ were 81 microM, 10 microM, 2.5 microM and 0.6 mM, respectively. The Km values for pyruvate, NADPH and bicarbonate were 4 mM, 25 microM and 20 mM, respectively. The enzyme converted malate to pyruvate (at pH 6.3) in the presence of 5 mM NAD+ at approximately 80 per cent of the maximum rate with NADP+. It exhibited oxaloacetate decarboxylase activity at about 10 per cent of the rate of oxidative decarboxylation of malate (with NADP+ as coenzyme) and pyruvate reductase activity at about 4 per cent of the rate of oxidative decarboxylation of malate with NADP+. The oxidative decarboxylation of malate was inhibited by malate at lower values of pH of incubation medium. This inhibition gradually decreased as the pH of the incubation medium increased. No inhibition was observed at pH 8.2. The addition of purified cytoplasmic malic enzyme, pyruvate, bicarbonate and NADPH generating system (consisting of NADP+, glucose 6-phosphate, glucose 6-phosphate dehydrogenase) stimulated about twofold progesterone biosynthesis by the isolated human placental mitochondria. This stimulation was abolished by arsenite and fluorocitrate. A possible role for the cytosolic malic enzyme in the regulation of progesterone biosynthesis in human placenta is discussed.

Electrophoresis, Polyacrylamide Gel↗