Fig. 2
Expression and purification of His-VP1 under denaturing conditions. A Soluble and pellet fractions of bacterial cells expressing His-VP1 analyzed on 10% SDS-PAGE. Lane 1 represents the protein markers, lane 2 and 3 represents the cellular protein fractions before and after IPTG induction, lane 4 represents the supernatant containing E.coli soluble proteins, and lane 5 represents the insoluble fraction containing His-VP1 B His-VP1 purified using nickel chelation chromatography under denaturing conditions analyzed on 10% SDS-PAGE. Lane 1 represents the protein markers, and lanes 2–8 represent protein fractions eluted with 350 mM imidazole. C Elution profile of His-GST-VP1 from a Superdex 200 (10/300) size exclusion column. D Purified protein from size exclusion chromatography analyzed on 10% SDS-PAGE. Lane 1 represents the protein markers, and lanes 2 and 3 represent the peak fraction. E Western blot showing the cross reactivity of the purified protein against an anti-HAV polyclonal antibody. F LC–MS/MS analysis showing mass spectra of trypsin digested VP1. G Peptide mass fingerprinting and database search showing the presence of HAV VP1