For
Escherichia coli, it has been assumed that
L-alanine is synthesized by alanine-valine transaminase (AvtA) in conjunction with an unknown alanine aminotransferase(s). We isolated alanine auxotrophs from a prototrophic double mutant deficient in AvtA and YfbQ, a novel alanine aminotransferase, by chemical mutagenesis. A shotgun cloning experiment identified two genes, uncharacterized
yfdZ and
serC, that complemented the alanine auxotrophy. When the
yfdZ- or
serC-mutation was introduced into the double mutant, one triple mutant (
avtA yfbQ yfdZ) showed alanine auxotrophy, and another (
avtA yfbQ serC), prototrophy. In addition, we found that four independent alanine auxotrophs possessed a point mutation in
yfdZ but not in
serC. We also found that
yfdZ expression was induced in minimal medium. Furthermore,
yfbQ-bearing plasmid conferred the ability to excrete alanine on the mutant lacking
D-amino acid dehydrogenase-encoding gene,
dadA. From these results, we concluded that
E. coli synthesizes
L-alanine by means of three aminotransferases, YfbQ, YfdZ, and AvtA.
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