title: Involvement of NarK1 and NarK2 proteins in transport of nitrate and nitrite in the denitrifying bacterium Pseudomonas aeruginosa PAO1.
abstract: Two transmembrane proteins were tentatively classified as NarK1 and NarK2 in the Pseudomonas genome project and hypothesized to play an important physiological role in nitrate/nitrite transport in Pseudomonas aeruginosa. The narK1 and narK2 genes are located in a cluster along with the structural genes for the nitrate reductase complex. Our studies indicate that the transcription of all these genes is initiated from a single promoter and that the gene complex narK1K2GHJI constitutes an operon. Utilizing an isogenic narK1 mutant, a narK2 mutant, and a narK1K2 double mutant, we explored their effect on growth under denitrifying conditions. While the DeltanarK1::Gm mutant was only slightly affected in its ability to grow under denitrification conditions, both the DeltanarK2::Gm and DeltanarK1K2::Gm mutants were found to be severely restricted in nitrate-dependent, anaerobic growth. All three strains demonstrated wild-type levels of nitrate reductase activity. Nitrate uptake by whole-cell suspensions demonstrated both the DeltanarK2::Gm and DeltanarK1K2::Gm mutants to have very low yet different nitrate uptake rates, while the DeltanarK1::Gm mutant exhibited wild-type levels of nitrate uptake. Finally, Escherichia coli narK rescued both the DeltanarK2::Gm and DeltanarK1K2::Gm mutants with respect to anaerobic respiratory growth. Our results indicate that only the NarK2 protein is required as a nitrate/nitrite transporter by Pseudomonas aeruginosa under denitrifying conditions.
authors.affiliation: Department of Biology, University of Dayton, 300 College Park, Dayton, OH 45469-2320, USA.
authors.forename: Vandana
authors.initials: V
authors.lastname: Sharma
authors.forename: Chris E
authors.initials: CE
authors.lastname: Noriega
authors.forename: John J
authors.initials: JJ
authors.lastname: Rowe
mesh_terms: D000693:Anaerobiosis; D027321:Anion Transport Proteins / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001693:Biological Transport, Active; D003470:Culture Media; D015964:Gene Expression Regulation, Bacterial; D005816:Genetic Complementation Test; D009154:Mutation; D000091285:Nitrate Transporters; D009566:Nitrates / Q000378:metabolism*; D009573:Nitrites / Q000378:metabolism*; D009876:Operon; D011550:Pseudomonas aeruginosa / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*
chemical_list: D027321:Anion Transport Proteins; D001426:Bacterial Proteins; D003470:Culture Media; D000091285:Nitrate Transporters; D009566:Nitrates; D009573:Nitrites