The Escherichia coli gabDTPC Operon: Specific γ-Aminobutyrate Catabolism and Nonspecific Induction

Abstract

Nitrogen limitation induces the nitrogen-regulated (Ntr) response, which includes proteins that assimilate ammonia and scavenge nitrogen. Nitrogen limitation also induces catabolic pathways that degrade four metabolically related compounds: putrescine, arginine, ornithine, and ␥-aminobutyrate (GABA). We analyzed the structure, function, and regulation of the gab operon, whose products degrade GABA, a proposed intermediate in putrescine catabolism. We showed that the gabDTPC gene cluster constitutes an operon based partially on coregulation of GabT and GabD activities and the polarity of an insertion in gabT on gabC. A ⌬gabDT mutant grew normally on all of the nitrogen sources tested except GABA. The unexpected growth with putrescine resulted from specific induction of gab-independent enzymes. Nac was required for gab transcription in vivo and in vitro. Ntr induction did not require GABA, but various nitrogen sources did not induce enzyme activity equally. A gabC (formerly ygaE) mutant grew faster with GABA and had elevated levels of gab operon products, which suggests that GabC is a repressor. GabC is proposed to reduce nitrogen source-specific modulation of expression. Unlike a wild-type strain, a gabC mutant utilized GABA as a carbon source and such growth required S . Previous studies showing S -dependent gab expression in stationary phase involved gabC mutants, which suggests that such expression does not occur in wild-type strains. The seemingly narrow catabolic function of the gab operon is contrasted with the nonspecific (nitrogen source-independent) induction. We propose that the gab operon and the Ntr response itself contribute to putrescine and polyamine homeostasis.

Ammonium/methylammonium transport (Amt) proteins facilitate diffusion of NH 3 bidirectionally

“…Degradation of arginine by the arginine succinyltransferase (Ast) catabolic pathway, the one present in enteric bacteria, yields 2 mol ammonium and 2 mol glutamate per mol of arginine (13,14). Our attempts to detect trace amounts of ammonium released into the medium by wild-type (amtB ϩ ) strains of S. typhimurium failed and hence we deduce that its concentration did not reach our detection limit of 20 M. To obtain further evidence that the product being released during growth on arginine was ammonium, we examined the ability of a wild-type strain to crossfeed a mutant strain (SK3512) that was unable to catabolize arginine because it carried a deletion of the entire arginine catabolic (ast) operon.…”

“…Progeny were initially selected on LB plates containing chloramphenicol and kanamycin (25 g͞ml). Plasmid pJES1293 was obtained from the shuttle vector pNo T a ͞T 7 by ligation of a 325-bp PCR fragment that carries the 5 prime end of the astC gene (first gene of the ast operon) (13,14). Plasmid pJES1294 was obtained from pNo T a ͞T 7 by ligation of a 311-bp PCR fragment that carries the 3 prime end of the astE gene (last gene of the ast operon).…”

“…Although many reports propose that Amt and methylammonium͞ammonium permease (MEP) proteins actively transport the charged species NH 4 ϩ (4, 5), there are several lines of evidence against this proposal (6,7). First, apparent concentration of the ammonium analog, [ 14 ϩ ), in an Amt͞MEP-dependent manner, was accounted for by metabolic trapping in enteric bacteria and by diffusion trapping into vacuoles and other acidic compartments in fungi. In each case accumulation depended on a subsequent energy-requiring reaction, one catalyzed by glutamine synthetase in bacteria or the V-type H ϩ -ATPase in fungi.…”

A Novel α-Ketoglutaric Semialdehyde Dehydrogenase

“…6A). Succinylglutamic semialdehyde dehydrogenase catalyzes the oxidization of succinylglutamic semialdehyde to succinylglutamate in the arginine succinyltransferase pathway of arginine degradation (43) and also acts on succinic semialdehyde (44). Indeed, SSALDH is involved in the alternative arginine decarboxylase pathway of arginine degradation.…”

Transcriptome Analysis of Agmatine and Putrescine Catabolism inPseudomonas aeruginosaPAO1

“…coli (30). In P. aeruginosa PAO1, the induction of gabDT by exogenous agmatine and putrescine was first revealed by GeneChip analyses (Table 2).…”