Genome analysis of Pseudomonas aeruginosa by field inversion gel electrophoresis

Grothues, D

doi:10.1016/0378-1097(87)90086-3

Cited by 28 publications

(34 citation statements)

References 9 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…P. aeruginosa PAO1 was encapsulated in agarose plugs as described by Grothues and Tümmler (25), but half of their bacterial cell concentration was used (5 ϫ 10 9 cells per ml). Single restriction digests with SpeI and DpnI were performed as described previously (55)(56)(57).…”

Section: Aminoacylated Trnamentioning

confidence: 99%

Regulation of the hemA gene during 5-aminolevulinic acid formation in Pseudomonas aeruginosa

et al. 1995

View full text Add to dashboard Cite

The general tetrapyrrole precursor 5-aminolevulinic acid is formed in bacteria via two different biosynthetic pathways. Members of the ␣ group of the proteobacteria use 5-aminolevulinic acid synthase for the condensation of succinyl-coenzyme A and glycine, while other bacteria utilize a two-step pathway from aminoacylated tRNA Glu . The tRNA-dependent pathway, involving the enzymes glutamyl-tRNA reductase (encoded by hemA) and glutamate-1-semialdehyde-2,1-aminomutase (encoded by hemL), was demonstrated to be used by Pseudomonas aeruginosa, Pseudomonas putida, Pseudomonas stutzeri, Comamonas testosteroni, Azotobacter vinelandii, and Acinetobacter calcoaceticus. To study the regulation of the pathway, the glutamyl-tRNA reductase gene (hemA) from P. aeruginosa was cloned by complementation of an Escherichia coli hemA mutant. The hemA gene was mapped to the SpeI A fragment and the DpnIL fragment of the P. aeruginosa chromosome corresponding to min 24.1 to 26.8. The cloned hemA gene, coding for a protein of 423 amino acids with a calculated molecular mass of 46,234 Da, forms an operon with the gene for protein release factor 1 (prf1). This translational factor mediates the termination of the protein chain at the ribosome at amber and ochre codons. Since the cloned hemA gene did not possess one of the appropriate stop codons, an autoregulatory mechanism such as that postulated for the enterobacterial system was ruled out. Three open reading frames of unknown function transcribed in the opposite direction to the hemA gene were found. hemM/orf1 and orf2 were found to be homologous to open reading frames located in the 5 region of enterobacterial hemA genes. While orf2 was found to be 59% identical to its enterobacterial counterpart, hemM/orf1 showed only 23% identity. The third open reading frame (orf3), located between the hemA gene and hemM/orf1, encodes a polypeptide with homology to outer membrane proteins. Biochemical and genetic evidence which makes a direct involvement of HemM/ Orf1 in the 5-aminolevulinic acid formation of P. aeruginosa very unlikely was obtained. An increase of hemA mRNA was observed under anaerobic denitrifying conditions, while anaerobic growth utilizing the fermentative arginine deiminase pathway led to a drastic decrease of hemA transcription compared with that observed for aerobically grown P. aeruginosa. These results suggest the anaerobic induction of hemA by the presence of nitrate. Two transcription start sites were located in the 5 region of the hemA gene. Utilization of both transcription start sites was changed in a P. aeruginosa mutant missing the oxygen regulator Anr (Fnr analog), indicating the involvement of the transcription factor in hemA expression. DNA sequences homologous to one half of an Anr binding site were detected at one of the determined transcription start sites.The tetrapyrrole content of a bacterial cell varies drastically depending on the conditions of the growth environment. The formation of 5-aminolevulinic acid (ALA), the first committed precursor of all tetra...

show abstract

Section: Aminoacylated Trnamentioning

confidence: 99%

Regulation of the hemA gene during 5-aminolevulinic acid formation in Pseudomonas aeruginosa

et al. 1995

View full text Add to dashboard Cite

show abstract

“…32 Agarose blocks were equilibrated in restriction enzyme buffer, and the chromosomal DNA embedded in the agarase was digested with either Xba I or Sfil. A slice of each insert plug was then sealed into a weil of 4-mm-thick horizontal agarase gel and electrophoresed (CHEF DR II, Bio-Rad Laboratories, München, Germany) in 0.5xTris-borate-EDTA buffer 49 for 25 h at 14aC and 200 V, with the pulse times increasing from 1 0 to 40 s. Lambda concatemers (Pharmacia, Freiburg, Germany), yeast chromosomes (S. cerevisiae WAY 5-4A, Biometra, Göttingen, Germany) and Hindill cleaved Iambda DNA were used as size markers.…”

Section: Sos-pagementioning

confidence: 99%

Clonal analysis of Escherichia coli serotype O6 strains from urinary tract infections

Zingler

Ott

Blum

et al. 1992

Microbial Pathogenesis

View full text Add to dashboard Cite

combinations of capsule ( K) and flagellin ( H) antigens, were analysed according to the outer membrane pattern (OMP), serum resistance properties, mannose-resistant hemagglutination using various types of erythrocytes, and also for the genetic presence and the expression of Pfimbriae. S fimbriae/F1 C fimbriae, Type 1 fimbriae, aerobactin and hemolysin. Twenty selected strains were further analysed by pulsed field gel electrophoresis (PFGE), elaborating genomic profilas by Xba I cleavage and subsequent Southern hybridization to virulence-associated DNA probes. lt could be shown that 06 UTI isolates represent a highly heterogeneaus group of strains according to the occurrence and combination of these traits. Relatedness an the genetic and the phenotypic Ievei was found for some of the strains exhibiting the same 0: K: H: F serotype. DNA Iang-range mapping further indicated some interesting features, according to the copy number and the genomic linkage of virulence genes.

show abstract

“…20 Agarose blocks containing genomic DNA were equilibrated in restriction enzyme buffer for 3 h on ice and cleaved in fresh buffer with 30 units restriction enzyme for 3 h at the appropriate incubation temperature. Orthogonal field alternation gel electrophoresis was performed with a Consort pulse field system, which includes a computer equipped E 654 power supply and a submarine basin (Consort, Belgium), using 1% agarase gels in 0.25 xTBE buffer.…”

Section: Orthogonal Field Alternation Gel Electrophoresis (Ofage)mentioning

confidence: 99%

Virulence determinants of Escherichia coli O6 extraintestinal isolates analysed by Southern hybridizations and DNA long range mapping techniques

Blum

Ott

Cross

et al. 1991

Microbial Pathogenesis

View full text Add to dashboard Cite

Various factors contribute to the virulence of these extraintestinal E. coli strains.Hemolysin production, expression of different adhesins and the synthesis of iron uptake substances such as aerobactin, as weil as the 0 and K surface antigens are important for the extraintestinal pathogenicity of E. coli. -6Fimbrial adhesins can be distinguished by differences in receptor specificities, which are detectable in vitro by the use of different target cells in agglutination assays.7 Type I fimbriae recognize mannose residues present e.g. on Saccharomyces cerevisiae cells. 8P fimbrial adhesins mediating adhesion to galactose-galactoside structures can be determined by using blood group P erythrocytes.

show abstract

Genome analysis of Pseudomonas aeruginosa by field inversion gel electrophoresis

Cited by 28 publications

References 9 publications

Regulation of the hemA gene during 5-aminolevulinic acid formation in Pseudomonas aeruginosa

Regulation of the hemA gene during 5-aminolevulinic acid formation in Pseudomonas aeruginosa

Clonal analysis of Escherichia coli serotype O6 strains from urinary tract infections

Virulence determinants of Escherichia coli O6 extraintestinal isolates analysed by Southern hybridizations and DNA long range mapping techniques

Contact Info

Product

Resources

About