Rotavirus VP7 epitope mapping using fragments of VP7 displayed on phages.
Vaccine
; 18(21): 2257-65, 2000 Apr 28.
Article
em En
| MEDLINE
| ID: mdl-10717346
ABSTRACT
cDNA copies of the complete porcine rotavirus CRW-8 VP7 gene were randomly digested to fragments of about 30-60 or 30-500 base pairs by DNase1 in the presence of Mn(2+). The fragments were cloned and expressed in a filamentous phage fd-tet-derived vector to create specific-gene-related peptide libraries. Polyclonal antibodies were then used to pan the SGRP libraries for antibody-binding phages. Analysis of the phage isolates revealed that the majority (86%) of them only had a single insert. However, phages displaying composite inserts containing the VP7 antigenic regions A, B, and C, originally defined by neutralising monoclonal antibody escape mutants, were also isolated. Inserts containing A or C region peptide were found to contain extra sequences from the C region, while the B region epitope was linear and had additional sequence from either upstream or downstream. In addition a dominant and possibly non-neutralising VP7 epitope was identified around amino acids 263-270. One of the recreated antigenic epitopes has also been fused to the outer membrane protein A (OmpA) of Escherichia coli and shown to maintain its antigenicity. The results in this study may have significant implication for recreation of conformational epitopes and vaccine development.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Fragmentos de Peptídeos
/
Bacteriófagos
/
Capsídeo
/
Rotavirus
/
Mapeamento de Epitopos
/
Proteínas do Capsídeo
/
Antígenos Virais
Limite:
Animals
Idioma:
En
Revista:
Vaccine
Ano de publicação:
2000
Tipo de documento:
Article
País de afiliação:
Austrália