Please use this identifier to cite or link to this item: https://doi.org/10.1016/j.jviromet.2011.10.006
Title: Monoclonal antibodies against dengue NS2B and NS3 proteins for the study of protein interactions in the flaviviral replication complex
Authors: Moreland, N.J.
Tay, M.Y.F.
Lim, E.
Rathore, A.P.S. 
Lim, A.P.C.
Hanson, B.J.
Vasudevan, S.G. 
Keywords: Dengue
Flavivirus
Monoclonal antibody
NS2B
NS3
Issue Date: Jan-2012
Citation: Moreland, N.J., Tay, M.Y.F., Lim, E., Rathore, A.P.S., Lim, A.P.C., Hanson, B.J., Vasudevan, S.G. (2012-01). Monoclonal antibodies against dengue NS2B and NS3 proteins for the study of protein interactions in the flaviviral replication complex. Journal of Virological Methods 179 (1) : 97-103. ScholarBank@NUS Repository. https://doi.org/10.1016/j.jviromet.2011.10.006
Abstract: The replication of dengue virus (DENV) RNA requires at least two viral non-structural (NS) proteins, NS3 and NS5. To facilitate the study of the DENV replication complex, human monoclonal IgG that are specific for NS proteins have been generated and characterised. The anti-NS3 IgG, 3F8, binds a conserved epitope (aa526-531) in the NS3 helicase domain, and cross-reacts with NS3 from all four DENV serotypes and the related yellow fever virus. The anti-NS2B IgG, 3F10, binds aa49-66 of NS2B (CF18), which forms part of the 47 aa hydrophilic cofactor region required for NS3 protease activity. The specificity of the IgG for their respective non-structural proteins has been demonstrated by immunofluorescence of cells infected with DENV and Western blotting. 3F8 is able to co-immunoprecipitate NS3 and NS5 from BHK-21 cells infected with DENV2, and 3F10 is able to detect an interaction between recombinant NS2B CF18NS3 full-length protein and the NS5 RNA-dependent RNA polymerase (RdRp) domain in an ELISA-based binding assay. The assay is specific and highly reproducible, with a clear binding curve seen when RdRp is incubated with increasing amounts of full-length NS3, but not the NS3 protease domain. The NS3 helicase domain competes with NS3 full-length for NS5 RdRp binding, with a K d. of 2.5μM. Since NS3 and NS5 are required for DENV replication, this fascile assay could be used to screen for non-nucleoside, allosteric inhibitors that disrupt the interaction between the two proteins. © 2011 Elsevier B.V.
Source Title: Journal of Virological Methods
URI: http://scholarbank.nus.edu.sg/handle/10635/110586
ISSN: 01660934
DOI: 10.1016/j.jviromet.2011.10.006
Appears in Collections:Staff Publications

Show full item record
Files in This Item:
There are no files associated with this item.

SCOPUSTM   
Citations

15
checked on May 27, 2018

WEB OF SCIENCETM
Citations

13
checked on Apr 10, 2018

Page view(s)

28
checked on May 11, 2018

Google ScholarTM

Check

Altmetric


Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.