Transposon mutations in the flagella biosynthetic pathway of the solvent-tolerant Pseudomonas putida S12 result in a decreased expression of solvent efflux genes.
Fourteen solvent-sensitive transposon mutants were generated from the solvent-tolerant Pseudomonas putida strain S12 by applying the TnMod-KmO mutagenesis system. These mutants were unable to grow in the presence of octanol and toluene. By cloning the region flanking the transposon insertion point a partial sequence of the interrupted genes was determined. Comparison of the deduced amino acid sequences with a protein database revealed the following interrupted putative gene products: organic solvent efflux proteins SrpA and SrpB, the flagellar structural proteins FlgK, FlaG, FliI, FliC, and FliH, the transcriptional activator FleQ, the alternative RNA polymerase sigma factor RpoN, and the flagellum-specific RNA polymerase sigma factor FliA (RpoF). The transposon mutants, except for the organic solvent efflux mutants, were nonmotile as determined by a swarm assay and the formation of the flagellum was totally impaired. Expression studies with a srp promoter probe showed a decreased expression of the SrpABC efflux pump in the nonmotile mutants.
Kieboom
J
J
Department of Food  Technology and Nutritional Sciences, Wageningen Agricultural University, The Netherlands. kieboom@bio.vu.nl
Bruinenberg
R
R
Keizer-Gunnink
I
I
de Bont
J A
JA
D001426:Bacterial Proteins / Q000235:genetics*; D002352:Carrier Proteins / Q000235:genetics; D003001:Cloning, Molecular; D005407:Flagella / Q000235:genetics* / Q000648:ultrastructure; D005408:Flagellin / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects / Q000502:physiology*; D026901:Membrane Transport Proteins; D016254:Mutagenesis, Insertional; D000442:Octanols / Q000378:metabolism; D009842:Oligopeptides; D010455:Peptides / Q000235:genetics; D006180:Proton-Translocating ATPases; D016958:Pseudomonas putida / Q000235:genetics* / Q000254:growth & development / Q000648:ultrastructure; D011994:Recombinant Proteins / Q000096:biosynthesis; D012808:Sigma Factor / Q000235:genetics; D012997:Solvents / Q000378:metabolism / Q000494:pharmacology; D014050:Toluene / Q000378:metabolism
D001426:Bacterial Proteins; D002352:Carrier Proteins; C088595:FlgK protein, Bacteria; C064971:FliA protein, Bacteria; D026901:Membrane Transport Proteins; D000442:Octanols; D009842:Oligopeptides; D010455:Peptides; D011994:Recombinant Proteins; D012808:Sigma Factor; D012997:Solvents; C109912:SrpA protein, Pseudomonas; C109913:SrpB protein, Pseudomonas putida; D005408:Flagellin; C069250:fliH protein, Bacteria; C069251:fliI protein, bacteria; D014050:Toluene; C085706:FLAG peptide; D006180:Proton-Translocating ATPases