Design of a chimeric protein composed of FimH, FyuA and CNF-1 virulence factors from uropathogenic Escherichia coli and evaluation its biological activity and immunogenicity in vitro and in vivo

• Published in: Microbial pathogenesis 2023-01, Vol.174, p.105920-105920, Article 105920
• Main Authors: Hedayat, Sheida, Habibi, Mehri, Hosseini Doust, Reza, Asadi Karam, Mohammad Reza
• Format: Article
• Language: English
• Subjects: Adhesins, Escherichia coli - genetics; Adjuvant; Animals; Bioinformatics' studies; Escherichia coli Infections - microbiology; Fimbriae Proteins; Humans; Immunoglobulin G; Mice; Molecular Docking Simulation; Multi-epitope; Rabbits; Recombinant Fusion Proteins - genetics; Urinary tract infection; Urinary Tract Infections - microbiology; Uropathogenic Escherichia coli; Uropathogenic Escherichia coli - genetics; Vaccine candidate; Virulence Factors - genetics
• ISSN: 0882-4010; 1096-1208
• DOI: 10.1016/j.micpath.2022.105920

Urinary tract infections (UTIs) caused by Uropathogenic Escherichia coli (UPEC) are among the most prevalent bacterial infections in humans. Antibiotic resistance among UPEC isolates is increasing, and designing an effective vaccine can prevent or reduce these infections. FimH adhesin, iron scavenger receptor FyuA, and cytotoxic necrotizing factor −1 (CNF-1) are among the most important virulence factors of UPEC strains. Thus, a novel multi-epitope protein composed of FimH, FyuA, and CNF-1 was designed to evaluate its biological activity and immunogenicity in vitro and in vivo, respectively. The final vaccine design had seven domains, including the N-terminal domain of FimH, four domains of FyuA, and two domains of CNF-1, as determined by immunoinformatics analysis. The results of tertiary structure prediction showed that the chimeric protein had a C-score of −0.25 and Z-score of −1.94. Molecular docking indicated that thirty six ligand residues of the chimeric protein interacted with 53 receptor residues of TLR-4 by hydrogen bonds and hydrophobic interactions. Analysis of protein expression by SDS-PAGE showed an approximately 44 kDa band with different concentrations of IPTG which were confirmed by Western blot. According to ELISA results, the level of IL-8 produced by stimulated Ht29 cells with the chimeric protein was significantly higher than the stimulated Ht29 cells with CNF-1 alone and un-stimulated Ht29 cells. Rabbits subcutaneously immunized with the chimeric protein admixed with Freund adjuvant induced higher level of serum IgG on day 14 after the first vaccination than control rabbits. Furthermore, the booster dose of the chimeric protein significantly enhanced the IgG levels as compared to day 14 and also controls. As, the chimeric protein has suitable B-cell epitopes and MHC-I and MHC-II binding epitopes to stimulate humoral and cellular immunity, it could be a promising vaccine candidate against UTIs caused by UPEC. Evaluating the multi-epitope protein in inducing humoral and cellular immune responses, as well as protection, is ongoing in the mice models. •We developed multi-epitope protein based on FimH, FyuA and CNF-1 of UPEC strain.•The protein has B-cell, MHC-I and MHC-II epitopes to stimulate different immunity arms.•IL-8 produced by stimulated Ht29 with the chimeric protein was higher than controls.•The chimeric protein could significantly increase IgG responses than control rabbits.•Multi-epitope vaccine FimH.FyuA.CNF-1 could be a prom