Novel HN1D4-4D9 monoclonal antibody neutralizes PPRV by blocking HN interaction with SLAM and nectin-4 receptors

Peste des petits ruminants (PPR) is a highly contagious viral disease devastating small ruminant livestock, posing a significant threat to global food security. While effective vaccines exist, antiviral therapeutics are lacking. The hemagglutinin-neuraminidase (HN) glycoprotein of Peste des petits ruminants virus (PPRV) is crucial for viral entry and is the primary target for protective humoral immunity. Understanding the functional antigenic determinants of the PPRV HN protein is vital for developing improved vaccines and targeted antiviral strategies.

Researchers developed a novel monoclonal antibody (mAb), HN1D4-4D9, to characterize functional epitopes on the PPRV HN protein. They performed epitope mapping analysis, including alanine-scanning mutagenesis, to identify key residues. Surface plasmon resonance was used to assess binding affinity. The neutralizing activity of HN1D4-4D9 was evaluated using virus pre-treatment, virus-antibody mixture, and post-attachment neutralizing assays in vitro. Mechanistic insights into viral adsorption were gained through quantitative real-time PCR and competitive co-immunoprecipitation assays to investigate HN interaction with host receptors.

Epitope mapping revealed that HN1D4-4D9 recognized a novel, conserved epitope (380ECLVEACK387), encompassing both a linear epitope (381CLVEACK387) and a conformational epitope (380ECLVEA385), consistent across multiple PPRV genotypes. Alanine-scanning mutagenesis identified several amino acid residues within this epitope as critical. Surface plasmon resonance confirmed the high-affinity binding of the identified HN epitope peptide to HN1D4-4D9. In vitro assays demonstrated that HN1D4-4D9 exhibited potent neutralizing activity when present during or before viral exposure. However, the antibody was ineffective after virus binding to cells, indicating its action at the viral adsorption stage. Mechanistically, quantitative real-time PCR and competitive co-immunoprecipitation assays revealed that HN1D4-4D9 blocks viral adsorption by specifically preventing the PPRV HN protein from interacting with host receptors SLAM and nectin-4. This blockade occurred without interfering with the fusion protein. Further biochemical assays confirmed that specific amino acid residues within the novel epitope are critical for interaction with both SLAM and nectin-4 receptors.

HN1D4-4D9 potently neutralizes PPRV by blocking viral adsorption through direct interference with the PPRV HN protein's interaction with host SLAM and nectin-4 receptors.