Epitope mapping of the non-structural protein 2 (NSP2) of rotavirus

Rotavirus infection accounts for millions of diarrhoeal cases among young children worldwide and severe diarrhoea can cause death following excessive dehydration. To date, numerous studies have been carried out to understand the pathogenesis and host immune reactions of rotavirus disease. As a result, a number of rotavirus vaccines have been developed to prevent the disease. In general, most studies have revealed that structural proteins, especially VP7 and VP4, elicit immune responses in humans following rotavirus infection. Similarly, a number of previous studies have also revealed animal and human antibody reactions targeted towards the non-structural proteins, including NSP2. However, little is known about the specific epitope region of NSP2 that is responsible for such antigenic reactions. This study was therefore designed to map the epitope region of NSP2 reacting to an NSP2-specific monoclonal antibody. Recombinant NSP2 proteins obtained through expression in an Escherichia coli expression system were first cleaved by 2-(2-nitrophenyl)-3 methyl-3-bromoindolenine (BNPS-skatole) and the resultant cleavage products were analysed by Western blotting using an anti-SA11 monoclonal antibody. Western blot results together with further dot blot, mass spectrometry and bioinformatics analyses revealed two putative epitope sequences on NSP2: QYNKFAVITHGKGHYRIVK and YRIVKYSSVANHADRVYAT. Nuclear magnetic resonance (NMR) studies further indicated that YNKF contributed to the antigenicity of the mapped epitope. Knowledge on the epitopes of NSP2 protein can be applied in further immunological studies of NSP2 and may be important in the future design of next generation rotavirus vaccines.