Expression and secretion of the cloned Pseudomonas aeruginosa exotoxin A by Escherichia coli.
AUTOR(ES)
Lory, S
RESUMO
The exotoxin A gene from Pseudomonas aeruginosa PAK was expressed in Escherichia coli from recombinant plasmids when transcription was initiated from a promoter in the cloning vector. The exotoxin A polypeptide synthesized was found to have an electrophoretic mobility in sodium dodecyl sulfate-polyacrylamide gels of 66,000 daltons, identical in size to the mature exotoxin A made by P. aeruginosa. Analysis of the location of exotoxin A in various bacterial compartments by immunoblotting revealed that exotoxin A was exported by E. coli into its periplasmic space. Several functional assays, including analyses of disulfide bond formation, potentiation of ADP-ribosyltransferase activity, and HeLa cell cytotoxicity, were used to establish that the conformation of exotoxin A isolated from the E. coli periplasmic space is identical to that of exotoxin exported by P. aeruginosa to its extracellular space. Previous studies with recombinant plasmids expressing exotoxin A from P. aeruginosa PA103 (G. D. Gray, D. Smith, J. Baldridge, R. Markins, M. Vasil, E. Chen, and M. Heyneker, Proc. Natl. Acad. Sci. USA 81:2645-2649, 1984) showed a complete lack of processing and export of pre-exotoxin A in E. coli, differing from results reported here. These discrepancies may be explained by observed differences in the sequence of signal peptides encoded by the exotoxin A genes of PAK and PA103 strains of P. aeruginosa.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=210713Documentos Relacionados
- Exotoxin A of Pseudomonas aeruginosa: active, cloned toxin is secreted into the periplasmic space of Escherichia coli.
- Expression of the argF gene of Pseudomonas aeruginosa in Pseudomonas aeruginosa, Pseudomonas putida, and Escherichia coli.
- Secretion of toxin A from Pseudomonas aeruginosa PAO1, PAK, and PA103 by Escherichia coli.
- Role of domain II of Pseudomonas exotoxin in the secretion of proteins into the periplasm and medium by Escherichia coli.
- Cloning and expression of Pseudomonas aeruginosa flagellin in Escherichia coli.
