Defining of MAbs-neutralizing sites on the surface glycoproteins Gn and Gc of a hantavirus using vesicular stomatitis virus pseudotypes and site-directed mutagenesis. (9th January 2019)

• Record Type: Journal Article
• Authors: Levanov, Lev; Iheozor-Ejiofor, Rommel Paneth; Lundkvist, Åke; Vapalahti, Olli; Plyusnin, Alexander
• Is Part Of: Journal of general virology. Volume 100:Number 2(2019)
• Page Start: 145
• Page End: 155
• Publication Date: 2019-01-09
• Subjects: Monoclonal antibody -- neutralizing epitope -- site-directed mutagenesis.; Virology -- Periodicals; Viruses; Microbiology; Virology; Virologie -- Périodiques; Microbiologie -- Périodiques; Virology; Virologie; Virologie; Electronic journals; Periodical; Periodicals; 579.2
• DOI: 10.1099/jgv.0.001202
• Languages: English
• ISSNs: 0022-1317

Abstract

Abstract : Earlier four monoclonal antibodies (MAbs) against surface glycoproteins Gn and Gc of puumala virus (PUUV, genus Orthohantavirus, family Hantaviridae, order Bunyavirales ) were generated and for three MAbs with neutralizing capacity the localization of binding epitopes was predicted using pepscan and phage-display techniques. In this work, we produced vesicular stomatitis virus (VSV) particles pseudotyped with the Gn and Gc glycoproteins of PUUV and applied site-directed mutagenesis to dissect the structure of neutralizing epitopes. Replacement of cysteine amino acid (aa) residues with alanines resulted in pseudotype particles with diminished (16 to 18 %) neut-titres; double Cys→Ala mutants, as well as mutants with bulky aromatic and charged residues replaced with alanines, have shown even stronger reduction in neut-titres (from 25 % to the escape phenotype). In silico modelling of the neut-epitopes supported the hypothesis that these critical residues are located on the surface of viral glycoprotein molecules and thus can be recognized by the antibodies indeed. A similar pattern was observed in experiments with mutant pseudotypes and sera collected from patients suggesting that these neut-epitopes are utilized in a course of human PUUV infection. These data will help understanding the mechanisms of hantavirus neutralization and assist construction of vaccine candidates.