title: Characterization of the intracellular glutamate decarboxylase system: analysis of its function, transcription, and role in the acid resistance of various strains of Listeria monocytogenes.
abstract: The glutamate decarboxylase (GAD) system is important for the acid resistance of Listeria monocytogenes. We previously showed that under acidic conditions, glutamate (Glt)/γ-aminobutyrate (GABA) antiport is impaired in minimal media but not in rich ones, like brain heart infusion. Here we demonstrate that this behavior is more complex and it is subject to strain and medium variation. Despite the impaired Glt/GABA antiport, cells accumulate intracellular GABA (GABA(i)) as a standard response against acid in any medium, and this occurs in all strains tested. Since these systems can occur independently of one another, we refer to them as the extracellular (GAD(e)) and intracellular (GAD(i)) systems. We show here that GAD(i) contributes to acid resistance since in a ΔgadD1D2 mutant, reduced GABA(i) accumulation coincided with a 3.2-log-unit reduction in survival at pH 3.0 compared to that of wild-type strain LO28. Among 20 different strains, the GAD(i) system was found to remove 23.11% ± 18.87% of the protons removed by the overall GAD system. Furthermore, the GAD(i) system is activated at milder pH values (4.5 to 5.0) than the GAD(e) system (pH 4.0 to 4.5), suggesting that GAD(i) is the more responsive of the two and the first line of defense against acid. Through functional genomics, we found a major role for GadD2 in the function of GAD(i), while that of GadD1 was minor. Furthermore, the transcription of the gad genes in three common reference strains (10403S, LO28, and EGD-e) during an acid challenge correlated well with their relative acid sensitivity. No transcriptional upregulation of the gadT2D2 operon, which is the most important component of the GAD system, was observed, while gadD3 transcription was the highest among all gad genes in all strains. In this study, we present a revised model for the function of the GAD system and highlight the important role of GAD(i) in the acid resistance of L. monocytogenes.
authors.affiliation: Bacterial Stress Response Group, Microbiology, School of Natural Sciences, National University of Ireland, Galway, Galway, Ireland. karatzas@hotmail.co.uk
authors.forename: Kimon-Andreas G
authors.initials: KA
authors.lastname: Karatzas
authors.forename: Laura
authors.initials: L
authors.lastname: Suur
authors.forename: Conor P
authors.initials: CP
authors.lastname: O'Byrne
mesh_terms: D000143:Acids / Q000378:metabolism* / Q000633:toxicity; D003470:Culture Media / Q000737:chemistry; D024881:Drug Resistance, Bacterial; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial; D005968:Glutamate Decarboxylase / Q000235:genetics* / Q000378:metabolism*; D008089:Listeria monocytogenes / Q000201:enzymology* / Q000235:genetics*; D050296:Microbial Viability / Q000187:drug effects; D013312:Stress, Physiological; D014158:Transcription, Genetic; D005680:gamma-Aminobutyric Acid / Q000378:metabolism
chemical_list: D000143:Acids; D003470:Culture Media; D005680:gamma-Aminobutyric Acid; D005968:Glutamate Decarboxylase