Comparative enzymology of (2S,4R)4-fluoroglutamine and (2S,4R)4-fluoroglutamate

Many cancer cells have a strong requirement for glutamine. As an aid for understanding this phenomenon the F-18-labeled 2S,4R stereoisomer of 4-fluoroglutamine [(2S,4R)4-FGln] was previously developed for in vivo positron emission tomography (PET). In the present work, comparative enzymological studies of unlabeled (2S,4R)4-FGln and its deamidated product (2S,4R)4-FGlu were conducted as an adjunct to these PET studies. Our findings are as follows: Rat kidney preparations catalyze the deamidation of (2S,4R)4FGln. (2,4R)4-FGln and (2S,4R)4-FGlu are substrates of various aminotransferases. (2S,4R)4-FGlu is a substrate of glutamate dehydrogenase, but not of sheep brain glutamine synthetase. The compound is, however, a strong inhibitor of this enzyme. Rat liver cytosolic fractions catalyze a gamma-elimination reaction with (2S,4R)4-FGlu, generating alpha-ketoglutarate. Coupling of a deamidase reaction with this gamma-elimination reaction provides an explanation for the previous detection of F-18(-) in tumors exposed to [F-18](2S,4R)4-FGIn. One enzyme contributing to this reaction was identified as alanine aminotransferase, which catalyzes competing gamma-elimination and aminotransferase reactions with (2S,4R)4-FGIu. This appears to be the first description of an aminotransferase catalyzing a gamma-elimination reaction. The present results demonstrate that (2S,4R)4FGIn and (2S,4R)4-FGIu are useful analogues for comparative studies of various glutamine- and glutamateutilizing enzymes in normal and cancerous mammalian tissues, and suggest that tumors may metabolize (2S,4R)4-FGIn in a generally similar fashion to glutamine. In plants, yeast and bacteria a major route for ammonia assimilation involves the consecutive action of glutamate synthase plus glutamine synthetase (glutamate synthase cycle). It is suggested that (2S,4R)4-FGIn and (2S,4R)4-FGIu will be useful probes in studies of ammonia assimilation by the glutamate synthase pathway in these organisms. Finally, glutamine transaminases are conserved in mammals, plants and bacteria, and probably serve to close the methionine salvage pathway, thus linking nitrogen metabolism to sulfur metabolism and one-carbon metabolism. It is suggested that (2S,4R)4-FGIn may be useful in studies of the methionine salvage pathway in a variety of organisms. (C) 2012 Elsevier Inc. All rights reserved.