Expression and characterization of uropathogenic Escherichia coli adhesin protein linked to cholera toxin A2B subunits in Escherichia coli TB1

The FimH subunit of type 1-fimbriated Escherichia coli (E. coli) has been determined as a major cause for urinary tract infections. Thus, to produce a possible vaccine antigen against urinary tract infections, the fimH gene was genetically coupled to the ctxa2b gene and cloned into a pMAL-p2E expression vector. The chimeric construction of pMALfimH/ctxa2b was then transformed into E. coli K-12 TB1 and its nucleotide sequence was verified. A fusion protein, based on fusing adhesin to the cholera toxin subunit A2B (CTXA2B), was induced with 0.01 mM isopropyl-β-D-thiogalactoside (IPTG) for 4 h at 37°C to yield a soluble fusion protein. The fusion protein was then purified by affinity chromatography. The expressed fusion protein was confirmed by SDS-PAGE and Western blotting using antibodies to the maltose binding protein (MBP) or the cholera toxin subunit B (CTXB), plus the N-terminal amino acid sequence was also analyzed. The orderly-assembled fusion protein was confirmed by a modified G_MI-ganglioside ELISA, using antibodies to adhesin. The results indicated that the purified fusion protein was an adhesin/CTXA2B protein containing E. coli adhesin and the G_MI-ganglioside binding activity of CTXB. Accordingly, this adhesin/CTXA2B protein may be a potential antigen for oral immunization against uropathogenic E. coli.