Lectin activity of Pseudomonas aeruginosa vaccine candidates PSE17-1, PSE41-5 and PSE54.

Pseudomonas aeruginosa is an opportunistic pathogen that causes nosocomial infections most commonly in immunocompromised, cystic fibrosis (CF) and burns patients. The pilin and Pseudomonas lectins 1 (PA-IL) and 2 (PA-IIL) are known glycan-binding proteins of P. aeruginosa that are involved in adherence to host cells, particularly CF host airways. Recently, new P. aeruginosa surface proteins were identified by reverse vaccinology and tested in vivo as potential vaccine antigens. Three of these, namely PSE17-1, PSE41-5 and PSE54, were screened for glycan binding using glycan arrays displaying glycan structures representative of those found on human cells. Surface plasmon resonance was used to confirm the lectin activity of these proteins, and determined affinities with several host glycans to be in the nanomolar range. PSE17-1 binds hyaluronic acid and sialyl Lewis A and X. PSE41-5 binds terminal ß-linked galactose structures, Lewis and ABO blood group antigens. PSE54 binds to ABO blood group antigens and some terminal ß-linked galactose. All three proteins are novel lectins of P. aeruginosa with potential roles in infection of host cells.

[Construction of MyD88-Pseudomonas aeruginosa epitope DNA vaccine and its expression in eukaryotic cells].

AIM: To construct the MyD88-Pseudomonas aeruginosa epitope vaccine and study its expression in eukaryotic cells. METHODS: To design and synthesize an epigene containing three B cell epitopes of OprF and one foreign "promiscuous" T cell epitope by overlapping extension PCR. tPA signal encoding sequence was amplified by PCR and then it was inserted into the 5' terminus of the epigene to construct tPA-OprF. tPA-OprF and MyD88 were cloned into the expression vector pIRES and the recombinant plasmid pIRES-tPAOprF-MyD88 was constructed. The recombinant plasmid was transfected into COS-7 cells by electroporation. The expression protein of tPA-OprF and MyD88 was detected by Western blot. RESULTS: The recombinant plasmid pIRES-tPA-OprF-MyD88 was successfully constructed. Western blot analysis indicated the tPA-OprF fusion protein was expressed in supertanant of COS-7 cells and MyD88 protein in COS-7 cells. CONCLUSION: The recombinant plasmid pIRES-tPA-OprF-MyD88 has been successfully constructed and tPA-OprF and MyD88 protein can be highly expressed in transfected cells. It may be used as a potential candidate of preventive vaccine of pseudomonas aeruginosa.