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Localization of antigenic sites of the E2 glycoprotein of transmissible gastroenteritis coronavirus
I Correa1, F Gebauer, M J Bullido
1Centro de Biología Molecular, CSIC-UAM, Facultad de Ciencias, Universidad Autónoma, Madrid, Spain.
Abstract:
Four antigenic sites of the E2 glycoprotein of transmissible gastroenteritis virus were defined by competitive radioimmunoassays of monoclonal antibodies (MAbs). Here, we describe the localization of these sites by testing the antigenicity of protein fragments and prokaryotic expression products of E2 gene fragments, and by sequencing of MAb-resistant (mar) mutants. Partial proteolysis of purified E2 protein allowed the isolation of a 28K fragment recognized by both site A- and site C-specific MAbs. An antiserum against this fragment bound to a synthetic peptide containing residues 1 to 18 and to an expression product containing residues 1 to 325. The same expression product was recognized by site C-specific MAbs. These data indicate that residues within the sequence 1 to 325 contribute to site C and possibly also to site A. Sequencing of mar mutants that escaped neutralization by site A-specific MAbs indicated that residues 538 and 543 also belong to site A. The binding of site-specific MAbs to expression products led directly to the localization of sites B and D, between residues 1 to 325 and 379 to 529, respectively. The first 37% of the polypeptide chain of E2 appears to be more immunogenic than the rest of the sequence.
Insights
Researchers mapped four antigenic sites on the E2 glycoprotein of transmissible gastroenteritis virus using monoclonal antibodies (MAbs). This study localized these sites on the E2 protein, revealing key regions for viral antigenicity.
Area of Science:
- Virology
- Immunology
- Molecular Biology
Background:
- Transmissible gastroenteritis virus (TGEV) poses a significant threat to swine health.
- The E2 glycoprotein is a major surface antigen of TGEV, crucial for viral entry and immune response.
- Understanding TGEV E2 antigenic sites is vital for vaccine development and antiviral strategies.
Purpose of the Study:
- To precisely localize the four defined antigenic sites on the TGEV E2 glycoprotein.
- To identify specific amino acid residues contributing to each antigenic site.
- To correlate antigenicity with specific regions of the E2 polypeptide chain.
Main Methods:
- Competitive radioimmunoassays using monoclonal antibodies (MAbs).
- Analysis of protein fragments generated by partial proteolysis.
- Characterization of prokaryotic expression products of E2 gene fragments.
- Sequencing of MAb-resistant (mar) mutants.
Main Results:
- Four distinct antigenic sites (A, B, C, D) were mapped on the TGEV E2 glycoprotein.
- Site C and potentially Site A involve residues within the 1-325 sequence.
- Residues 538 and 543 were identified as part of Site A.
- Site B localized to residues 1-325, and Site D to residues 379-529.
- The initial 37% of the E2 polypeptide chain demonstrated higher immunogenicity.
Conclusions:
- The study successfully localized TGEV E2 antigenic sites, providing a detailed map of MAb-binding regions.
- Specific amino acid sequences within E2 were identified as critical for antibody recognition.
- These findings offer valuable insights for rational vaccine design targeting TGEV.