SUMMARYA 7000 Mr cleavage fragment of the F 1 subunit that carries the major neutralization epitope has been identified by chemical and enzymatic cleavage of the fusion protein of respiratory syncytial (RS) virus (Long strain) with an efficient RS virus-neutralizing monoclonal antibody. Based on the published mRNA-deduced sequence of the A2 strain, coupled to the hydropathicity profile and prediction of protein conformation, the neutralization epitope has tentatively been localized on the first third of the F1 protein N-terminal, probably in the region of amino acids 215Ser to 236Glu. Analysis of three peptides covering different portions of the 212Cys to 236Glu region of the F 1 fusion protein identified a peptide (Cys. 216Ash to 236Glu) that reacted strongly with the neutralizing monoclonal antibody and that was efficient in blocking neutralization and in plaque-reducing assays, confirming that the neutralization epitope was localized in that region. Further analysis with two other synthetic peptides (212Cys to 222Glu and Cys.221Ile to 236Glu) indicated that the dodecapeptide Ile-Glu-Phe-Gln-LysAsn-Asn-Arg-Leu-Leu-Glu mimicked either the whole or a major part of the neutralization epitope. This opens a promising avenue for the simple design of a synthetic peptide vaccine to control RS virus infection. INTRODUCTIONThe genus Pneumovirus of the Paramyxoviridae family consists of human and bovine strains of respiratory syncytial (RS) virus and pneumonia virus of mice. Human RS virus, an important pathogen in young children (Belshe et al., 1984), is thought to be a monotypic strain though minor differences in cross-neutralization assays have been noted. Using monoclonal antibodies (MAbs), Mufson et al. (1985) have been able to separate different isolates into subgroups A and B; however, studies by Prince et al. (1985) suggest that antigenic differences detected in vitro disappear when virus strains are compared in vivo. Recently, using a neutralizing MAb produced against the Long strain of RS virus, a major and highly conserved neutralization epitope was identified which was present on human, bovine and caprine strains of RS virus but not on pneumonia virus of mice. This epitope is localized on the F 1 fragment of the fusion glycoprotein (Trudel et al., 1986(Trudel et al., , 1987.The fusion glycoprotein is found as a 125K Mr dimer on the viral envelope and is involved in cell fusion leading to the formation of syncytia and cell penetration. It is composed of two disulphide-linked subunits: F1 with a reported Mr varying from 43K to 50K and F2 from 19K to 25K (Walsh et al., 1985, 1986;Trudel et al., 1986).Expression by recombinant vaccinia viruses of the major and fusion glycoproteins of RS virus (Olmsted et al., 1986) confirmed previous work carried out with MAbs and purified solubilized proteins showing the predominant role of the fusion glycoprotein in inducing immunity
Rubella virus (RV)-specific immunoglobulin G antibodies were studied by enzyme-linked immunosorbent assay (ELISA) techniques in sera from RV (RA 27/3)-vaccinated individuals, patients experiencing natural RV infection, congenital rubella syndrome patients, and individuals failing to respond to repeated RV immunization. Results obtained by using whole-RV ELISAs (detergent-solubilized M33 strain or intact Gilchrist strain) and hemagglutination inhibition (HAI) and neutralization (NT) assays were compared with results obtained with the same sera by using ELISAs employing a synthetic peptide, BCH-178, representing a putative neutralization domain on the RV El protein. Murine RV El-specific monoclonal antibodies with HAI and NT activities exhibited strong reactivity in ELISAs with BCH-178 peptide. In sera from RA 27/3-vaccinated individuals collected at 0 (prevaccine), 1, 2, 3, 4, 5, 6, 12, and 24 to 52 weeks postvaccine, the development of El-peptide-reactive antibodies closely paralleled increases in RV-specific antibodies measured by whole-RV ELISAs and HAI and NT assays. Similarly, sequential serum samples obtained from patients during acute and convalescent phases of natural RV infection showed a coordinate increase in RV-specific antibodies as measured by whole-RV and peptide ELISAs. Conversely, congenital rubella syndrome patient sera, although exhibiting high levels of antibody in whole-RV ELISAs, had little or no antibody directed to the neutralization domain peptide. Sera from patients failing to respond to repeated RV immunization contained very low levels of RV-specific antibody in all ELISAs. Our results suggest that the sequence represented by BCH-178 peptide may be a previously unidentified neutralization epitope for human antibodies on the RV El protein and may prove useful in determining effective RV immunity.
We have previously reported that rat Sertoli cells in culture produce and secrete plasminogen activator, a highly specific protease, and that FSH stimulates these processes.
A surface probability method was used to select a decapeptide homologous to residues 993 to 1002 of the peplomer protein E2 of murine hepatitis virus strain JHM, a neurotropic coronavirus. This sequence of amino acids corresponded to a minor peak on a hydrophilicity plot. Immunization of mice with the chemically synthesized peptide coupled to keyhole limpet hemocyanin elicited high levels of neutralizing antibody and protected against lethal virus challenge. Protection correlated with a critical level of antipeptide antibody, which could be reached after a single inoculation. These results suggest that an appropriate antibody response to a highly restricted, surface-exposed domain of this viral protein is critical in determining the outcome of infection of the central nervous system. This sequence is located in the C-terminal fifth of the E2 peplomers, between two predicted coiled-coil structures.
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