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 s ynthetic peptide could be cleaved by trypsin and therefore possessed t he 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 re action of anti-peptide serum with infectious virus neutralized infecti vity thereby supporting the biological importance of this site. Anothe r interesting characteristic of this peptide was its ability to bind t o the nucleocapsid protein resulting in a laddering effect on the nucl eocapsid 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, 5866 (1985); Sabara et al., J. Gen. Virol. 67, 201-212 (1986)]. Defin itive proof of binding was provided by the fact that the increments in the ladder corresponded to the molecular weight of the synthetic pept ide and that anti-peptide serum specifically reacted with the ladder f ormations. The laddering of the nucleocapsid could be eliminated by in cubation with trypsin thus further supporting the formation of a synth etic peptide-nucleocapsid complex. Due to the ability of the peptide t o bind to trypsin and to the nucleocapsid protein its biological activ ity was investigated. It appeared that increasing concentrations of th e peptide reduced the rate of virus plaque formation, thereby suggesti ng 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 abili ty to complex tightly to another protein. In protection-challenge expe riments performed using a murine model, animals immunized with VP4-pep tide provided protection passively, to neonates suckling on the immune darns, against a virulent rotavirus. The potential applications of th is peptide in rotavirus diagnosis, therapy and synthetic peptides base d vaccine is discussed.