Characterization of a synthetic peptide mimicking trypsin-cleavage site of rotavirus VP4.

A synthetic peptide corresponding to the trypsin cleavage site on the 84 k protein of bovine rotavirus was synthesized (VP4-peptide). This synthetic peptide could be cleaved by trypsin and therefore possessed the enzyme binding site present on the authentic protein. Further proof that this peptide mimicks the authentic trypsin cleavage site was the specific reaction of anti-peptide serum with the 84 k protein. The reaction of anti-peptide serum with infectious virus neutralized infectivity thereby supporting the biological importance of this site. Another interesting characteristic of this peptide was its ability to bind to the nucleocapsid protein resulting in a laddering effect on the nucleocapsid monomer (45 k), dimer (90 k) and trimer (135 k) [Gorzilia et al., J. Gen. Virol. 66, 1889-1900 (1985); Sabara et al., J. Virol. 53, 58-66 (1985); Sabara et al., J. Gen. Virol. 67, 201-212 (1986)]. Definitive proof of binding was provided by the fact that the increments in the ladder corresponded to the molecular weight of the synthetic peptide and that anti-peptide serum specifically reacted with the ladder formations. The laddering of the nucleocapsid could be eliminated by incubation with trypsin thus further supporting the formation of a synthetic peptide-nucleocapsid complex. Due to the ability of the peptide to bind to trypsin and to the nucleocapsid protein its biological activity was investigated. It appeared that increasing concentrations of the peptide reduced the rate of virus plaque formation, thereby suggesting that virus replication was inhibited. These results illustrate two features of this synthetic peptide which warrant further investigation; (1) its capacity to mimic an enzyme cleavage site and, (2) its ability to complex tightly to another protein. In protection-challenge experiments performed using a murine model, animals immunized with VP4-peptide provided protection passively, to neonates suckling on the immune dams, against a virulent rotavirus. The potential applications of this peptide in rotavirus diagnosis, therapy and synthetic peptides based vaccine is discussed.