A Panel of Monoclonal Antibodies Targeting the Rabies Virus Phosphoprotein Identifies a Highly Variable Epitope of Value for Sensitive Strain Discrimination

Abstract

A recombinant rabies virus phosphoprotein fusion product (GST-P) was used to generate a series of monoclonal antibodies (MAbs) with anti-P reactivity. Competitive binding assays classified 27 of these MAbs into four groups (I to IV), and 24 of them were deemed to recognize linear epitopes, as judged by their reaction in immunoblots. The linear epitope recognized in each case was mapped by using two series of N- and C-terminally deleted recombinant phosphoproteins. Assessment of the reactivities of representative MAbs to a variety of lyssavirus isolates by an indirect fluorescent antibody test indicated that group I MAbs, which recognized a highly conserved N-terminal epitope, were broadly cross-reactive with all lyssaviruses assayed, while group III MAbs, which reacted with a site overlapping that of group I MAbs, exhibited variable reactivities and group IV MAbs reacted with most isolates of genotypes 1, 6, and 7 only. In contrast, group II MAbs, which recognized an epitope located within a highly divergent central portion of the protein, were exquisitely strain specific. These anti-P MAbs are potentially useful tools for lyssavirus identification and discrimination.

Lyssavirus P Gene Characterisation Provides Insights into the Phylogeny of the Genus and Identifies Structural Similarities and Diversity within the Encoded Phosphoprotein

“…Similarly the poorly conserved VD2 might function as a spacer/hinge segment analogous to the hinge region of VSV P located between two functionally important domains (Banerjee and Barik, 1992). Indeed, epitope mapping studies of several anti-RABV P monoclonal antibodies raised against the ARCTIC strain have demonstrated the accessibility and immunogenicity of an exquisitely strain-specific antigenic site (AS II) within VD2 (Nadin-Davis et al, 2000), properties consistent with a surface structure of limited conservation. However, a panel of anti-MOKV P monoclonal antibodies completely lacked representatives of AS II (S. Nadin-Davis, unpublished data).…”

“…The CD1 region, which includes the N-terminal 20 amino acids proposed to contribute N protein (Fu et al, 1994) and L protein (Chenik et al, 1998) binding sites, is clearly under substantial structural constraint, a feature which appears to extend to the N-terminal 50 aa segment previously shown to contain an antigenic site (AS I) widely conserved in the Lyssavirus genus (Nadin-Davis et al, 2000). The functional significance of CD2 is less clearly defined apart from its contribution, via a lysinerich motif, to the strong N-binding activity which also requires the leucine-rich, hydrophobic 50 amino acid segment at the protein's C-terminal (Fu et al, 1994;Chenik et al, 1994;Jacob et al, 2001), a region which may be functionally equivalent to domain III of VSV P.…”

“…Schematic illustrating the organization of the lyssavirus P protein. Antigenic sites have been defined using anti-P monoclonal antibodies directed against the RABV (Nadin-Davis et al, 2000) and MOKV P proteins (unpublished data). The positions of the five serine residues targeted for phosphorylation in the CVS strain are denoted 1-5 and indicated by solid lines (conserved) or dashed lines (less conserved).…”

Raiva: uma breve revisão

“…O primeiro método laboratorial rápido proposto para o diagnóstico de raiva foi a detecção de corpúsculos de Negri, método este descrito por Adelchi Negri há mais de um século [90,104]. Negri acreditava que estas inclusões eram formas de um protozoário que ele julgava ser o causador da raiva.…”

Development and evaluation of an anti-rabies virus phosphoprotein-specific monoclonal antibody for detection of rabies neutralizing antibodies using RFFIT

“…In addition, although only RABV belonging to genotype 1 has been reported in Korea to date, bat species that are potential hosts of bat-related lyssavirus inhabit Korea, thereby posing an additional risk. Since P protein contains a broadly cross-reactive epitope for all lyssaviruses, monoclonal antibodies (mAbs) targeting P protein have been proven to be potentially useful for diagnosis and serotyping [ 9 ]. To this end, in this study, we developed an Alexa-conjugated monoclonal antibody (mAb) that specifically combines with P protein, using the first domestic KGH strain isolated from a human.…”