Aim. The aim is obtaining, investigation and selection of recombinant antigens for inclusion theirs into the against Pseudomonas vaccine. Materials and methods. The genes encoding of the outer membrane proteins F, L and I and Exotoxin A were synthesized by PCR with the genomic DNA of Pseudomonas aeruginosa. The amplified sequences were cloned into plasmid vectors for expression in cells of Escherichia coli. As the result of expression were the synthesized recombinant proteins that were purified in columns with a nickel-activated sorbent. The authenticity of the recombinant antigens was assessed by electrophoresis and immunoblotting. For assessing the immunogenicity of the recombinant proteins,they were sorbed on aluminum hydroxide and used for intraperitoneal immunization of mice. After a course of immunization, mice were injected intraperitoneally with a live virulent culture or еxotoxin A. Results. The obtained recombinant outer membrane proteins OprF, OprL and OprI, as well as the deletion variant of еxotoxin A (toxoid) stimulated immune reactions and protected the experimental animals from the virulent culture of P. aeruginosa. Using of the complexes of the recombinant proteins, as well as immunization with the fusion proteins consisting from sequences of two or three recombinant antigens, produced an additive increase in protective effects. The combination of the recombinant OprF protein and the recombinant toxoid (efficiency index of protective properties (EI 3.0) and two recombinant fusion proteins (EI 3.5) were the most effective. The first recombinant fusion protein (OprF-aTox-OprI) consisted from fused polypeptide sequences of OprF, toxoid and OprI. The second recombinant fusion protein (OprF-OprI) consisted from fused polypeptide sequences of OprF and OprI. Conclusion. The data obtained showed the fundamental possibility of using recombinant fusion proteins OprF-aTox-OprI and OprF-OprI as well as the complex of the recombinant OprF protein and the recombinant toxoid as the candidated vaccines against Pseudomonas aeruginosa.
The influence of the ratio of protein and aluminum hydroxide on the manifestation of the protective properties of the recombinant fusion protein OprF-aTox-OprI Pseudomonas aeruginosa and the expression of genes of molecules of innate immunity
