Functional characterization of the dguRABC locus for D-Glu and d-Gln utilization in Pseudomonas aeruginosa PAO1

D-Glu, an essential component of peptidoglycans, can be utilized as a carbon and nitrogen source by Pseudomonas aeruginosa. DNA microarrays were employed to identify genes involved in D-Glu catabolism. Through gene knockout and growth phenotype analysis, the divergent dguR-dguABC (D-Glu utilization) gene cluster was shown to participate in D-Glu and D-Gln catabolism and regulation. Growth of the dguR and dguA mutants was abolished completely on D-Glu or retarded on D-Gln as the sole source of carbon and/or nitrogen. The dguA gene encoded a FAD-dependent D-amino acid dehydrogenase with d-Glu as its preferred substrate, and its promoter was specifically induced by exogenous D-Glu and D-Gln. The function of DguR as a transcriptional activator of the dguABC operon was demonstrated by promoter activity measurements in vivo and by mobility shift assays with purified His-tagged DguR in vitro. Although the DNA-binding activity of DguR did not require D-Glu, the presence of D-Glu, but not D-Gln, in the binding reaction was found to stabilize a preferred nucleoprotein complex. The presence of a putative DguR operator was revealed by in silica analysis of the dguR-dguA intergenic regions among Pseudomonas spp. and binding of DguR to a highly conserved 19 bp sequence motif was further demonstrated. The dguB gene encodes a putative enamine/imine deaminase of the RidA family, but its role in D-Glu catabolism remains to be determined. Whilst a lesion in dguC encoding a periplasmic solute binding protein only affected growth on D-Glu slightly, expression of the previously characterized AatJMQP transporter for acidic l-amino acid uptake was found inducible by D-Glu and essential for D-Glu utilization. In summary, the findings of this study supported DguA as a new member of the FAD-dependent d-amino acid dehydrogenase family, and DguR as a D-Glu sensor and transcriptional activator of the dguA promoter.