Bk. Janes et al., Growth inhibition caused by overexpression of the structural gene for glutamate dehydrogenase (gdhA) from Klebsiella aerogenes, J BACT, 183(8), 2001, pp. 2709-2714
Two linked mutations affecting glutamate dehydrogenase (GDH) formation (gdh
-1 and rev-2) had been isolated at a locus near the trp cluster in Klebsiel
la aerogenes. The properties of these two mutations were consistent with th
ose of a locus containing either a regulatory gene or a structural gene. Th
e gdhA gene from K. aerogenes was cloned and sequenced, and an insertion mu
tation was generated and shown to be linked to trp. A region of gdhA from a
strain bearing gdh-1 was sequenced and shown to have a single-base-pair ch
ange, confirming that the locus defined by gdh-1 is the structural gene for
GDH. Mutants with the same phenotype as rev-2 were isolated, and their seq
uences showed that the mutations were located in the promoter region of the
gdhA gene. The linkage of gdhA to trp in K. aerogenes was explained by pos
tulating an inversion of the genetic map relative to other enteric bacteria
. Strains that bore high-copy-number clones of gdhA displayed an auxotrophy
that was interpreted as a limitation for alpha -ketoglutarate and conseque
ntly for succinyl-coenzyme A (CoA). Three lines of evidence supported this
interpretation: high-copy number clones of the enzymatically inactive gdhA1
allele showed no auxotrophy, repression of GDH expression by the nitrogen
assimilation control protein (NAC) relieved the auxotrophy, and addition of
compounds that could increase the alpha -ketoglutarate supply or reduce th
e succinyl-CoA requirement relieved the auxotrophy.