[Production of hybrid protein OprF-OprL of Pseudomonas aeruginosa]

Abstract

Aim: Production of preparation consisting of amino acid sequences of 2 proteins of outer membrane--OprF and OprI--of P. aeruginosa and study of its protective properties from experimental P. aeruginosa infection.

Materials and methods: Nucleotide sequences coding OprF protein (1 kb) as well as its C-terminal region (0.6 kb) and OprI protein (0.25 kb) were integrated into pQE-30 plasmid (QIAGEN). And oprF gene (C-terminal region of oprF in variant 2) and oprI gene were combined and cloned sequentially into a single vector. E. coli M15 strain cells (QIAGEN) were used for the production of producent strains of recombinant proteins. Protein products were analyzed by electrophoresis in polyacrylamide gel by Lammle. Purification of recombinant proteins was performed by affinity chromatography in Ni-sepharose columns. Live virulent culture P. aeruginosa PA-170015 strain was used for the analysis of protective properties of recombinant proteins.

Results: 2 hybrid recombinant proteins were produced including amino acid sequences of F and I proteins of outer membrane (OprF and OprI) of P. aeruginosa. Recombinant protein 1 included whole size sequences of OprF and OprI and protein 2--C-terminal region (including amino acid residues 192-342) of OprF and whole size sequence of OprI. These recombinant proteins after 2 immunizations protected mice from the experimental intraperitoneal infection with P. aeruginosa. Hybrid protein consisting of whole size sequences had the best protective effect.

Conclusion: The results obtained open a perspective for further immunobiological testing of hybrid recombinant protein OprF-OprI with the aim of creating immunopreparations for prophylaxis of P. aeruginosa infection.