Purification and characterization of LasR as a DNA-binding protein.
In Pseudomonas aeruginosa, the activator protein LasR and a cognate autoinducer (AI) are required for expression of the elastase gene (lasB). In the present study, we investigated the binding properties of the P. aeruginosa lasR gene product. The LasR protein was overexpressed and purified as a glutathione S-transferase (GST) fusion protein. Using gel retardation and UV cross-linking analysis, we demonstrated that the GST-LasR could bind to a separate site in the lasB upstream operator regions 1 and 3 in the presence of the autoinducer. Regions 1 and 3 are located at 105 and 42 base pairs upstream, respectively, from the lasB transcriptional start site. Our present results clearly demonstrate that LasR is a specific DNA-binding protein that regulates the transcription of the lasB gene in the presence of an autoinducer.
You
Z
Z
Department of Bacteriology, Yokohama City University School of Medicine, Japan.
Fukushima
J
J
Ishiwata
T
T
Chang
B
B
Kurata
M
M
Kawamoto
S
S
Williams
P
P
Okuda
K
K
D015107:4-Butyrolactone / Q000031:analogs & derivatives / Q000378:metabolism; D001426:Bacterial Proteins; D001665:Binding Sites; D015153:Blotting, Western; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D005822:Genetic Vectors / Q000235:genetics; D005982:Glutathione Transferase / Q000235:genetics; D008666:Metalloendopeptidases / Q000235:genetics / Q000378:metabolism; D010957:Plasmids; D011550:Pseudomonas aeruginosa / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C080365:LasR protein, Pseudomonas aeruginosa; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; C088386:homoserine lactone; D005982:Glutathione Transferase; D008666:Metalloendopeptidases; C043859:pseudolysin, Pseudomonas aeruginosa; D015107:4-Butyrolactone