The conversion of alanine into glutamine in guinea-pig renal cortex. Essential role of pyruvate carboxylase

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Abstract
The metabolism of L-alanine was studied in isolated guinea pig kidney-cortex tubules. In contrast with the conclusions of Krebs (1935) glutamine was the main carbon and nitrogenous product of the metabolism of alanine (at 1 and 5 mM). Glutamate and NH3 were only minor products. At neither concentration of alanine was there accumulation of glucose, glycogen, pyruvate, lactate, aspartate or tricarboxylic acid-cycle intermediates. C-balance calculations and the release of 14CO2 from [U-14C]alanine indicate that oxidation of the alanine C skeleton occurred at both substrate concentrations. A pathway involving alanine aminotransferase, glutamate dehydrogenase, glutamine synthetase, pyruvate dehydrogenase, pyruvate carboxylase and enzymes of the tricarboxylic acid cycle is proposed for the conversion of alanine into glutamine. Strong evidence for this pathway was obtained by: suppressing alanine removal by amino-oxyacetate, an inhibitor of transaminase; measuring the release of 14CO2 from [1-14C]alanine; the use of L-methionine DL-sulfoximine, an inhibitor of glutamine synthetase, which induced a large increase in ammonia release from alanine; and the use of fluoroacetate, an inhibitor of aconitase, which inhibited glutamine synthesis with concomitant accumulation of citrate from alanine. In this pathway the central role of pyruvate carboxylase, which explains the discrepancy between the results and those of Krebs (1935), was also demonstrated.