A quantitative analysis of the control of glutamine catabolism in rat liver cells. Use of selective inhibitors

Abstract

1. At a physiological concentration of glutamine (0.5 mM), 87% of the total transport across the plasma membrane of liver cells isolated from fed rats involved the Na(+)-dependent system N; this was substantially inhibited by L-histidine. The residual Na(+)-independent component was attributed to system L on the basis of inhibition by 2-amino-2-norbornanecarboxylate and L-tryptophan. 2. Catabolism of glutamine by intact liver cells or by isolated mitochondria was inhibited by glutamate gamma-hydrazide with IC50 values of 13.7 +/- 3.5 microM and 22.6 +/- 3.8 microM respectively and a maximal inhibition of approx. 75%. The site of inhibition was identified as glutaminase; glutamate gamma-hydrazide inhibited this enzyme in cell-free extracts (IC50 37.8 +/- 7.7 microM) but had no activity against glutamate dehydrogenase or transport of glutamine, whether across mitochondrial or plasma membranes. 3. The major control site in cells from fed animals incubated with 0.5 mM L-glutamine was glutaminase (flux control coefficient 0.96). Appreciable control also resided in both plasma membrane transport systems, with coefficients of 0.51 for system N and -0.46 for system L, such that both interacted to provide a fine control of the intracellular concentration of the amino acid. Similar values were obtained by computer simulation based on theoretical determination of elasticities. 4. Previous controversy about the locus of regulation of hepatic glutamine metabolism is resolved by this distribution of control.