Glutamate synthesis and the control of reactions linked with the nicotinamide-adenine dinucleotide coenzymes in mitochondria

Abstract

The rate of glutamate synthesis from citrate or a-oxoglutarate and ammonia by intact respiring guinea-pig liver mitochondria was studied under various conditions affecting the relative proportions of reduced and oxidized intramitochondrial [nicotinamide-adenine dinucleotide] (NAD) and [nicotinamide-adenine dinucleotide phosphate] (NADP). Rapid oxidation and reduction of NAD and NADP by respiration favor a high rate of glutamate synthesis. The rate of reoxidation of intramitochondrial NADH and NADPH on addition of ammonia in the. absence of respiration was shown fluorimefrically to be much more rapid when a-oxoglutarate was the substrate than when citrate was being oxidized. The activities of isocitrate dehydrogenase and glutamate dehydrogenase were studied in broken liver mitochondria. The rate of glutamate synthesis by such systems in the presence of isocitrate, ammonia and NADPH was compared with glutamate synthesis by intact mitochondria. Guinea-pig-mammary-gland mitochondria contain no detectable glutamate dehydrogenase; neither in the presence of citrate nor of a-oxoglutarate did the addition of ammonia stimulate the oxidation of NADH and NADPH or the synthesis of glutamate. The kinetics of the various phenomena studied can be interpreted in terms of delays in availability of NAD and NADP both to enzymes which reduce them and to systems which oxidize them, such as the respiratory chain or a special synthetic couple like glutamate dehydrogenase.