K-041. Characterization of Two ATP Phosphoribosyltransferase Isoenzymes of Geobacter sulfurreducens

M. A. Aklujkar, D. R. Lovley;
Univ. of Massachusetts, Amherst, MA.

Adenosine triphosphate (ATP) phosphoribosyltransferase (HisG), the first enzyme of the histidine biosynthesis pathway, is of two kinds: a longer protein that functions independently or a shorter protein that requires a regulatory subunit, HisZ. Geobacter sulfurreducens and related Fe(III)-respiring bacteria, which are of interest for their role in uranium bioremediation and ability to produce electricity, are unique in that they possess both types of hisG genes, and the hisZ gene resembles a full-length histidyl-tRNA synthetase gene rather than one that lacks the anticodon loop recognition domain. Deletion mutants of the long hisG-1, the short hisG-2, and hisZ were constructed and characterized. Deletion of hisG-2 greatly reduced the growth rate of G. sulfurreducens in the absence of exogenous histidine, whereas deletion of hisG-1 had no effect. A double deletion mutant was unable to grow unless histidine was added to the growth medium. These results indicate that HisG-2 provides the major ATP phosphoribosyltransferase activity of Geobacteraceae, but HisG-1 has sufficient activity to sustain some growth, and that Geobacteraceae can grow auxotrophically by uptake of histidine from the growth medium. Surprisingly, deletion of hisZ resulted in a prolonged growth lag regardless of histidine supplementation, followed by adaptation. This may be due to a polar effect of the deletion on the 3’ adjacent gene purA, encoding adenylosuccinate synthase, the first enzyme of the adenosine-specific branch of the purine biosynthesis pathway, resulting in an ATP shortage. Transcriptional and translational coupling of hisZ and purA is likely to ensure sufficient flux of 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR), to which ATP is degraded during histidine biosynthesis, towards adenosine rather than guanosine. The unprecedented redundancy of ATP phosphoribosyltransferases in Geobacteraceae suggests that an unidentified metabolite, such as a vitamin or a metal-binding compound, may be made from histidine or an intermediate of the histidine biosynthesis pathway, and regulate HisG-1 activity by feedback inhibition.