title: Characterization of epitopes for virus-neutralizing monoclonal antibodies to Ross River virus E2 using phage-displayed random peptide libraries.
abstract: Ross River virus (RRV) is the predominant cause of epidemic polyarthritis in Australia, yet the antigenic determinants are not well defined. We aimed to characterize epitope(s) on RRV-E2 for a panel of monoclonal antibodies (MAbs) that recognize overlapping conformational epitopes on the E2 envelope protein of RRV and that neutralize virus infection of cells in vitro. Phage-displayed random peptide libraries were probed with the MAbs T1E7, NB3C4, and T10C9 using solution-phase and solid-phase biopanning methods. The peptides VSIFPPA and KTAISPT were selected 15 and 6 times, respectively, by all three of the MAbs using solution-phase biopanning. The peptide LRLPPAP was selected 8 times by NB3C4 using solid-phase biopanning; this peptide shares a trio of amino acids with the peptide VSIFPPA. Phage that expressed the peptides VSIFPPA and LRLPPAP were reactive with T1E7 and/or NB3C4, and phage that expressed the peptides VSIFPPA, LRLPPAP, and KTAISPT partially inhibited the reactivity of T1E7 with RRV. The selected peptides resemble regions of RRV-E2 adjacent to sites mutated in neutralization escape variants of RRV derived by culture in the presence of these MAbs (E2 210-219 and 238-245) and an additional region of E2 172-182. Together these sites represent a conformational epitope of E2 that is informative of cellular contact sites on RRV.
authors.affiliation: Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria 3168, Australia. Janet.Davies@med.monash.edu.au
authors.forename: J M
authors.initials: JM
authors.lastname: Davies
authors.forename: Y P
authors.initials: YP
authors.lastname: Cai
authors.forename: R C
authors.initials: RC
authors.lastname: Weir
authors.forename: M J
authors.initials: MJ
authors.lastname: Rowley
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000914:Antibodies, Viral / Q000276:immunology*; D000918:Antibody Specificity / Q000276:immunology; D002213:Capsid / Q000737:chemistry / Q000276:immunology*; D036022:Capsid Proteins; D003001:Cloning, Molecular; D018604:Epitope Mapping; D000939:Epitopes / Q000737:chemistry / Q000276:immunology*; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D010446:Peptide Fragments / Q000737:chemistry / Q000276:immunology; D019151:Peptide Library; D010802:Phylogeny; D012398:Ross River virus / Q000276:immunology*; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein; D012995:Solubility; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000276:immunology*
chemical_list: D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D036022:Capsid Proteins; C085817:E2 protein, Ross River virus; D000939:Epitopes; D010446:Peptide Fragments; D019151:Peptide Library; D014759:Viral Envelope Proteins