The glutamine-depend acid resistance system in neutralophilic bacteria

Background: The amino acid-dependent acid resistance (AR) systems are widespread in neutralophilic bacteria. Their activities overlap so to cover a rather large acidic pH range, from 6 to <2. Recently, the AR system relying on glutamine (AR2_Q) was shown to be operative in Escherichia coli, Lactobacillus reuteri and some Brucella species. AR2_Q requires the glutaminase GlsA/YbaS and the membrane antiporter GadC, both proteins are active at acidic pH. GadC imports L-glutamine and exports either glutamate (the glutamine deamination product) or GABA, the decarboxylation product of glutamate, via glutamate decarboxylase (GadB), a structural component of the glutamate-dependent AR system, together with GadC (AR2). Objectives: This work was undertaken to study the distribution of the AR2_Q in bacteria and to develop a qualitative acid-glutaminase assay and a quantitative GadC-mediated transport assay. Methods: The qualitative assay takes advantage of the color change of the pH indicator bromocresol green. The quantitative assay is fluorometric, HPLC-based, to concomitantly measure glutamine uptake, and glutamate and GABA export. Results: The colorimetric assay is fast and reliable. It also provides information on co-occurrence of AR2 and AR2_Q in bacterial species. The HPLC-based assay provided evidence that glutamine uptake and the selective export of glutamate and/or GABA via GadC is finely modulated in the pH range 2.5-4.0. A bioinformatic genome analysis showed that the gene encoding acid-glutaminase is often nearby or in operon arrangement with the genes encoding GadC and/or GadB. Overall, our results suggest that AR2_Q is likely to be of prominent importance especially for enteric bacteria AR.