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Major receptor-binding and neutralization determinants are located within the same domain of the transmissible
M Godet1, J Grosclaude, B Delmas
1Unité de Virologie et Immunologie Moléculaires, Institut National de la Recherche Agronomique, Jouy-en-Josas, France.
Abstract:
The spike glycoprotein (S) of coronavirus, the major target for virus-neutralizing antibodies, is assumed to mediate the attachment of virions to the host cell. A 26-kilodalton fragment proteolytically cleaved from transmissible gastroenteritis virus (TGEV) S protein was previously shown to bear two adjacent antigenic sites, A and B, both defined by high-titer neutralizing antibodies. Recombinant baculoviruses expressing C-terminal truncations of the 26-kilodalton region were used to localize functionally important determinants in the S protein primary structure. Two overlapping 223- and 150-amino-acid-long products with serine 506 as a common N terminus expressed all of the site A and B epitopes and induced virus-binding antibodies. Coexpression of one of these truncated protein S derivatives with aminopeptidase N (APN), a cell surface molecule acting as a receptor for TGEV, led to the formation of a complex which could be immunoprecipitated by anti-S antibodies. These data provide evidence that major neutralization-mediating and receptor-binding determinants reside together within a domain of the S protein which behaves like an independent module. In spite of their ability to prevent S-APN interaction, the neutralizing antibodies appeared to recognize a preformed complex, thus indicating that antibody- and receptor-binding determinants should be essentially distinct. Together these findings bring new insight into the molecular mechanism of TGEV neutralization.
Insights
Transmissible gastroenteritis virus (TGEV) spike protein contains key neutralization and receptor-binding sites within a single domain. Neutralizing antibodies target distinct sites from the virus receptor, aminopeptidase N (APN).
Area of Science:
- Virology
- Immunology
- Molecular Biology
Background:
- The spike (S) glycoprotein of coronaviruses is crucial for host cell attachment and a primary target for neutralizing antibodies.
- A 26-kilodalton fragment of the transmissible gastroenteritis virus (TGEV) S protein contains two adjacent antigenic sites, A and B, recognized by neutralizing antibodies.
Purpose of the Study:
- To identify functionally important determinants within the TGEV S protein primary structure.
- To investigate the relationship between antibody-binding and receptor-binding sites on the TGEV S protein.
Main Methods:
- Utilized recombinant baculoviruses to express C-terminal truncations of the TGEV S protein's 26-kilodalton region.
- Coexpressed truncated S protein derivatives with aminopeptidase N (APN), the TGEV receptor.
- Employed immunoprecipitation to detect protein complexes.
Main Results:
- Two overlapping truncated S protein products (223- and 150-amino-acid) retained all site A and B epitopes and induced virus-binding antibodies.
- Coexpression with APN formed a complex immunoprecipitable by anti-S antibodies, indicating a distinct domain for receptor binding.
- Neutralizing antibodies, while blocking S-APN interaction, recognized a preformed complex, suggesting distinct antibody and receptor binding sites.
Conclusions:
- Major neutralization-mediating and receptor-binding determinants are localized within a single, modular domain of the TGEV S protein.
- Antibody and receptor binding sites on the TGEV S protein are essentially distinct, despite their proximity and functional roles.