The synthesis of hippurate from benzoate and glycine by rat liver mitochondria. Submitochondrial localization and kinetics.
1. Rat liver mitochondria make hippurate at up to 4 nmol/min per mg of protein. The rate of synthesis supported by oxidation of glutamate with exogenous Pi present is identical with that supported by ATP plus oligomycin. Lower rates were obtained with other respiratory substrates, and when glutamate was used without Pi. 2. A matrix localization for hippurate synthesis is indicated by the latency of benzoyl-CoA synthetase and glycine N-acyltransferase to their extramitochondrial substrates, failure of exogenous benzoyl-CoA to inhibit incorporation of [14C]hippurate and inhibition of hippurate synthesis supported by ATP, but not glutamate, by carboxyatractyloside. 3. The relative activities of the individual enzymes and the mitochondrial content of benzoyl-CoA in the presence and absence of glycine suggest that hippurate synthesis is rate-limited by formation of benzoyl-CoA. 4. The increases in rates of ATP hydrolysis and of O2 consumption on the addition of benzoate and glycine were in good agreement with those required to support hippurate synthesis. The increase in respiration indicates that State-4 respiration [Chance & Williams (1957) Adv. Enzymol 17, 65-134] is not used, with these conditions, for ATP synthesis.
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D000963:Antimetabolites / Q000494:pharmacology; D001565:Benzoates / Q000378:metabolism*; D002956:Citrulline / Q000096:biosynthesis; D003065:Coenzyme A / Q000378:metabolism; D005998:Glycine / Q000378:metabolism*; D006626:Hippurates / Q000096:biosynthesis*; D066298:In Vitro Techniques; D007700:Kinetics; D008297:Male; D008930:Mitochondria, Liver / Q000378:metabolism*; D010101:Oxygen Consumption; D051381:Rats
D000963:Antimetabolites; D001565:Benzoates; D006626:Hippurates; D002956:Citrulline; D000255:Adenosine Triphosphate; D003065:Coenzyme A; D005998:Glycine