title: Functional characterization of the dguRABC locus for D-Glu and d-Gln utilization in Pseudomonas aeruginosa PAO1.
abstract: 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.
authors.affiliation: Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, PR China.|Department of Biology, Georgia State University, Atlanta, Georgia, USA.
authors.forename: Weiqing
authors.initials: W
authors.lastname: He
authors.affiliation: Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, PR China.|Department of Biology, Georgia State University, Atlanta, Georgia, USA.
authors.forename: Guoqing
authors.initials: G
authors.lastname: Li
authors.affiliation: Department of Biology, Georgia State University, Atlanta, Georgia, USA.
authors.forename: Chun-Kai
authors.initials: CK
authors.lastname: Yang
authors.affiliation: Department of Biology, Georgia State University, Atlanta, Georgia, USA.
authors.forename: Chung-Dar
authors.initials: CD
authors.lastname: Lu
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D003605:D-Amino-Acid Oxidase / Q000235:genetics / Q000378:metabolism*; D055786:Gene Knockout Techniques; D023061:Gene Order; D056426:Genetic Loci; D018698:Glutamic Acid / Q000378:metabolism*; D005973:Glutamine / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism; D053858:Metabolic Networks and Pathways / Q000235:genetics*; D005810:Multigene Family; D011550:Pseudomonas aeruginosa / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D026901:Membrane Transport Proteins; D015534:Trans-Activators; D005973:Glutamine; D018698:Glutamic Acid; D003605:D-Amino-Acid Oxidase