Uropathogenic E.coli PapG Protein: The Expression and Immune Characterization in Balb/C Mice

Abstract

Urinary tract infection (UTI) is one of the most complications in human caused by uropathogenic Escherichia coli (UPEC) and there is no effective vaccine against UTI. In this study the PapG protein was expressed and monitored for immunoassay. The pEXA/PapG vector was digested with NdeI/EcoRI enzymes, and the gene segment was sub-cloned into pET21a vector. After transformation to E. coli strain Origami, PapG was expressed under the 1 mM IPTG. Protein expression was detected by SDS-PAGE and western blotting. Protein purification was carried out using Ni-NTA column. Experimental mice (n=18) were divided into three groups. The groups 1 to 3 were immunized three times with 2 weeks interval with the PapG protein with freund’s adjuvant, alum adjuvanted protein and PBS, as a control. Two weeks after the last injection, lymphocyte proliferation assessed with Brdu method, secretion of IL-4 and IFN-γ cytokines were quantified by ELISA kit. In addition, total antibody, IgG1 and IgG2a subtypes were assayed with ELISA. The results indicated that PapG, as a vaccine candidate, significantly increase lymphocyte proliferation and Th1 cytokine pattern versus control group. Humoral immune responses induced with both IgG1 and IgG2a classes. Although PapG, as a vaccine candidate, strongly induced cellular and humoral immune responses, more studies and experimental bacterial challenges remained to be clarified

Keywords

• Uropathogenic E.coli, PapG, expression, immunization, immune responses