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Pseudomonas aeruginosa  is an important pathogen causing urinary tract infections (UTIs) and resistance to antibiotics has increased the need for a vaccine against this bacterium.  P. aeruginosa  V-antigen (PcrV), which is a component of the type III secretion system, delivers exoenzymes such as exoenzyme S (ExoS) into the host cells. In the present study, we aimed to design and express a hybrid protein composed of PcrV and ExoS from  P. aeruginosa  using bioinformatics. Finally, pre-existing antibodies were evaluated in sera collected from patients with UTI. The prediction results showed that the hybrid protein ExoS.PcrV had a C-score of −0.85 and Z-score of −5.55 versus C-score of −2.93 and Z-score of −2.65 for PcrV.ExoS. Based on BepiPred and ABCpred, 15 and 14 linear B-cell epitopes, as well as five conformational epitopes were identified in ExoS.PcrV. The interaction between the protein and immune receptor was validated  in silico . Molecular docking indicated that the hybrid protein interacted strongly with Toll-like receptor 2. ExoS.PcrV was expressed in pET28a-BL21 and purified with a size of 57 kD by Nickel resins. The protein reacted with all sera collected from humans infected with  P. aeruginosa  following Western blot. The infected patients produced significantly higher IgG levels against the protein compared to the control as indicated by ELISA. The protein ExoS.PcrV was determined as a promising candidate against UTI caused by  P. aeruginosa  and the presence of pre-existing antibodies indicated the advantage of the hybrid protein. Evaluation of the efficacy of hybrid protein is ongoing in mice model. Communicated by Ramaswamy H. Sarma

journal of biomolecular structure and dynamics

Bioinformatics analyses for the designation of a hybrid protein against urinary tract infections caused by  Pseudomonas aeruginosa  and investigation of the presence of pre-existing antibodies in infected humans