Glutamate deamination by glutamate dehydrogenase plays a central role in amino acid catabolism in plants

Abstract

Glutamate is of central importance in plant N metabolism since the biosynthesis of all other amino acids requires this compound. Glutamate dehydrogenase (GDH; EC 1.4.1.2), which catalyzes in vitro reversible reductive amination of 2-oxoglutatre to form glutamate, is a key player in the metabolism of glutamate. While most previous studies have indicated that the oxidative deamination is the in vivo direction of the GDH reaction, its physiological role has remained ambiguous for decades. We have recently isolated mutants for the two known Arabidopsis GDH genes and created a gdh double mutant. Our recent work revealed an increased susceptibility of the gdh double mutant to dark-induced C starvation, the first phenotype associated with the loss of GDH activity in plants. Monitoring the amino acid breakdown during the dark treatment also suggested that the deamination of glutamate catalyzed by GDH is central to the catabolism of many other amino acids.Nitrogen is a major limiting factor for plant growth and development and therefore the incorporation of inorganic N into amino acids is of central importance in agriculture. GDH had been viewed as the major ammonium assimilatory enzyme in the past. However, the discovery of glutamine synthetase (GS)/glutamate synthase (GOGAT) pathway as the sole port of ammonium assimilation raised the long-lasting question regarding the role played by GDH. 1,2 This was confounded by the uncertainty about the in vivo direction of the GDH reaction, which reversibly aminates 2-oxoglutatrate or deaminates glutamate in vitro. Plants contain at least two GDH genes encoding either the α or β subunit of GDH. 3 Random assembling of the two subunits in the hexameric holoenzyme results in the seven isoenzymes which can be visualized on a non-denaturing gel [β 6 (isoenzyme 1), α 1 β 5 … α 5 β 1, α 6 (isoenzyme 7)]. 4 For the in vivo direction that GDH catalyzes, oxidative deamination has been supported by many labeling experiments. 2 In two recent studies, the overexpression of either α or β subunit of GDH resulted in the increased release of 15 NH 4 + after [ 15 N]glutamate feeding, demonstrating the deaminating role of the distinct GDH isoenzymes. 5,6 In contrast, 15 NH 4 + incorporation into glutamate through GDH was demonstrated under salt stress in which continued 15 NH 4 + assimilation occurred in the presence of the GS inhibitor, methionine sulfoximine. 7 Nevertheless, most previous results supported the current consensus that GDH deaminates glutamate in vivo under normal growth conditions.Although previous labeling experiments were useful to define the in vivo direction of the GDH reaction, the physiological role of GDH remained speculative. For this reason, GDH expression has been extensively studied at the transcript, protein and enzyme activity levels. In these previous studies, the regulation of GDH by light/dark cycle, sugar levels and ammonia addition was documented. 2 In brief, expression of GDH appears to be controlled by the cellular energy state or sugar levels since the dark-ind...

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