期刊
VIRUS RESEARCH
卷 179, 期 -, 页码 225-230出版社
ELSEVIER
DOI: 10.1016/j.virusres.2013.11.010
关键词
Dengue; NS5; Phage display; Antibody; Cross-reactivity
类别
资金
- DUKE-NUS Signature Research Program
- Ministry of Health, Singapore
- National Medical Research Council, Singapore [NMRC/1315/2011]
The multifunctional dengue nonstructural (NS) protein 5 from the four serotypes of dengue virus (DENV1-4) is essential for viral replication and harbors a methyl transferase (MTase) and a RNA-dependent RNA-polymerase domain (RdRp). There are limited comparative studies of NS5 from the four DENV serotypes and this is further hampered by a lack of cross-reactive NS5 antibodies. In this study, recombinant NS5 proteins were expressed, purified, enzymatically characterized, and used strategically as bait in biopanning experiments with a naive human Fab phage-display library to identify serotype specific or cross-reactive Fab fragments. Using a combination of peptide competition ELISA and peptide phage display the epitopes of the cross-reactive Fabs were mapped to the first alpha helix of the MTase domain (5M1) and the priming loop of the RdRp domain (5R3). The epitope of a third, serotype-specific Fab (5M3) was mapped to aa19-30 of the DENV3 MTase domain. Together the recombinant proteins and specific antibodies will facilitate further mechanistic studies of the DENV replication complex. (C) 2013 Elsevier B.V. All rights reserved.
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