A glycosylated peptide in the West Nile virus envelope protein is immunogenic during equine infection.
Abstract: Using a monoclonal antibody directed to domain I of the West Nile virus (WNV) envelope (E) protein, we identified a continuous (linear) epitope that was immunogenic during WNV infection of horses. Using synthetic peptides, this epitope was mapped to a 19 aa sequence (WN19: E147-165) encompassing the WNV NY99 E protein glycosylation site at position 154. The inability of WNV-positive horse and mouse sera to bind the synthetic peptides indicated that glycosylation was required for recognition of peptide WN19 by WNV-specific antibodies in sera. N-linked glycosylation of WN19 was achieved through expression of the peptide as a C-terminal fusion protein in mammalian cells and specific reactivity of WNV-positive horse sera to the glycosylated WN19 fusion protein was shown by Western blot. Additional sera collected from horses infected with Murray Valley encephalitis virus (MVEV), which is similarly glycosylated at position E154 and exhibits high sequence identity to WNV NY99 in this region, also recognized the recombinant peptide. Failure of most WNV- and MVEV-positive horse sera to recognize the epitope as a deglycosylated fusion protein confirmed that the N-linked glycan was important for antibody recognition of the peptide. Together, these results suggest that the induction of antibodies to the WN19 epitope during WNV infection of horses is generally associated with E protein glycosylation of the infecting viral strain.
Publication Date: 2008-11-15 PubMed ID: 19008394DOI: 10.1099/vir.0.2008/003731-0Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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This study focuses on understanding the West Nile virus (WNV), specifically a glycosylated peptide in the virus’s envelope protein that is immunogenic during infection in horses. The researchers found that the recognition of this peptide by WNV-specific antibodies in horse and mouse sera required glycosylation.
Identification of Immunogenic Epitope
- The researchers used a monoclonal antibody, which is an antibody produced by a single clone of cells, to target domain I of the WNV envelope protein. This allowed them to identify an immunogenic epitope, which is a part of an antigen that an immune cell recognizes and responds to. During WNV infection in horses, this epitope could trigger an immune response.
- Through the use of synthetic peptides, they mapped this epitope to a 19 amino acid sequence. This sequence included the WNV NY99 E protein glycosylation site at position 154. Glycosylation is the process where a carbohydrate is added to a protein, affecting its stability and function.
- The researchers found that WNV-positive horse and mouse sera (the clear liquid that can be separated from clotted blood) could not bind the synthetic peptides without glycosylation, showing the importance of this process in immune response.
N-linked Glycosylation and Reactivity
- The team achieved N-linked glycosylation of the peptide WN19 by expressing it as a C-terminal fusion protein in mammalian cells. They demonstrated that WNV-positive horse sera specifically reacted to the glycosylated WN19 fusion protein, confirmed through a Western blot test.
- Additionally, the researchers observed that sera from horses infected with Murray Valley encephalitis virus (MVEV), which is also glycosylated at position E154 and has a high sequence identity to WNV NY99 in this region, recognized the recombinant peptide.
- Most importantly, the study revealed that the failure of most WNV-positive and MVEV-positive horse sera to recognize the epitope as a deglycosylated fusion protein confirmed the importance of the N-linked glycan for antibody recognition of the peptide.
Implications for Antibody Induction in Horses
- The results of the study suggest a general association between the induction of antibodies to the WN19 epitope and the E protein glycosylation of the infecting viral strain during WNV infection of horses. In other words, the research provides insights into how horses’ immune systems respond to WNV infection and the crucial role glycosylation plays in these responses.
Cite This Article
APA
Hobson-Peters J, Toye P, Sánchez MD, Bossart KN, Wang LF, Clark DC, Cheah WY, Hall RA.
(2008).
A glycosylated peptide in the West Nile virus envelope protein is immunogenic during equine infection.
J Gen Virol, 89(Pt 12), 3063-3072.
https://doi.org/10.1099/vir.0.2008/003731-0 Publication
Researcher Affiliations
- Australian Biosecurity CRC for Emerging Infectious Disease, St Lucia, Queensland, Australia.
- AGEN Biomedical Limited, Acacia Ridge, Queensland, Australia.
- School of Molecular and Microbial Sciences, The University of Queensland, St Lucia, Queensland 4072, Australia.
- AGEN Biomedical Limited, Acacia Ridge, Queensland, Australia.
- Department of Microbiology, School of Medicine, University of Pennsylvania, Philadelphia, PA, USA.
- CSIRO Livestock Industries, Australian Animal Health Laboratory, Geelong, Victoria, Australia.
- Australian Biosecurity CRC for Emerging Infectious Disease, St Lucia, Queensland, Australia.
- CSIRO Livestock Industries, Australian Animal Health Laboratory, Geelong, Victoria, Australia.
- Australian Biosecurity CRC for Emerging Infectious Disease, St Lucia, Queensland, Australia.
- School of Molecular and Microbial Sciences, The University of Queensland, St Lucia, Queensland 4072, Australia.
- School of Molecular and Microbial Sciences, The University of Queensland, St Lucia, Queensland 4072, Australia.
- School of Molecular and Microbial Sciences, The University of Queensland, St Lucia, Queensland 4072, Australia.
MeSH Terms
- Amino Acid Sequence
- Animals
- Antibodies, Monoclonal / blood
- Antibodies, Monoclonal / immunology
- Antibodies, Viral / blood
- Antibodies, Viral / immunology
- COS Cells
- Cell Line
- Chlorocebus aethiops
- Encephalitis Virus, Murray Valley / immunology
- Epitope Mapping
- Glycosylation
- Horse Diseases / diagnosis
- Horse Diseases / immunology
- Horse Diseases / virology
- Horses
- Molecular Sequence Data
- Peptides / chemical synthesis
- Peptides / chemistry
- Peptides / immunology
- Vero Cells
- Viral Envelope Proteins / chemistry
- Viral Envelope Proteins / immunology
- West Nile Fever / diagnosis
- West Nile Fever / immunology
- West Nile Fever / veterinary
- West Nile Fever / virology
- West Nile virus / immunology
Citations
This article has been cited 11 times.- Plante JA, Plante KS, Popov VL, Shinde DP, Widen SG, Buenemann M, Nogueira ML, Vasilakis N. Morphologic and Genetic Characterization of Ilheus Virus, a Potential Emergent Flavivirus in the Americas. Viruses 2023 Jan 10;15(1).
- Hardy JM, Newton ND, Modhiran N, Scott CAP, Venugopal H, Vet LJ, Young PR, Hall RA, Hobson-Peters J, Coulibaly F, Watterson D. A unified route for flavivirus structures uncovers essential pocket factors conserved across pathogenic viruses. Nat Commun 2021 Jun 1;12(1):3266.
- Prow NA, Edmonds JH, Williams DT, Setoh YX, Bielefeldt-Ohmann H, Suen WW, Hobson-Peters J, van den Hurk AF, Pyke AT, Hall-Mendelin S, Northill JA, Johansen CA, Warrilow D, Wang J, Kirkland PD, Doggett S, Andrade CC, Brault AC, Khromykh AA, Hall RA. Virulence and Evolution of West Nile Virus, Australia, 1960-2012. Emerg Infect Dis 2016 Aug;22(8):1353-62.
- Goh LY, Hobson-Peters J, Prow NA, Baker K, Piyasena TB, Taylor CT, Rana A, Hastie ML, Gorman JJ, Hall RA. The Chikungunya Virus Capsid Protein Contains Linear B Cell Epitopes in the N- and C-Terminal Regions that are Dependent on an Intact C-Terminus for Antibody Recognition. Viruses 2015 Jun 8;7(6):2943-64.
- Hussmann KL, Vandergaast R, Zheng K, Hoover LI, Fredericksen BL. Structural proteins of West Nile virus are a major determinant of infectious particle production and fitness in astrocytes. J Gen Virol 2014 Sep;95(Pt 9):1991-2003.
- Prow NA. The changing epidemiology of Kunjin virus in Australia. Int J Environ Res Public Health 2013 Nov 25;10(12):6255-72.
- Prow NA, Tan CS, Wang W, Hobson-Peters J, Kidd L, Barton A, Wright J, Hall RA, Bielefeldt-Ohmann H. Natural exposure of horses to mosquito-borne flaviviruses in south-east Queensland, Australia. Int J Environ Res Public Health 2013 Sep 17;10(9):4432-43.
- Hobson-Peters J, Yam AW, Lu JW, Setoh YX, May FJ, Kurucz N, Walsh S, Prow NA, Davis SS, Weir R, Melville L, Hunt N, Webb RI, Blitvich BJ, Whelan P, Hall RA. A new insect-specific flavivirus from northern Australia suppresses replication of West Nile virus and Murray Valley encephalitis virus in co-infected mosquito cells. PLoS One 2013;8(2):e56534.
- Edmonds J, van Grinsven E, Prow N, Bosco-Lauth A, Brault AC, Bowen RA, Hall RA, Khromykh AA. A novel bacterium-free method for generation of flavivirus infectious DNA by circular polymerase extension reaction allows accurate recapitulation of viral heterogeneity. J Virol 2013 Feb;87(4):2367-72.
- Hobson-Peters J. Approaches for the development of rapid serological assays for surveillance and diagnosis of infections caused by zoonotic flaviviruses of the Japanese encephalitis virus serocomplex. J Biomed Biotechnol 2012;2012:379738.
- Frost MJ, Zhang J, Edmonds JH, Prow NA, Gu X, Davis R, Hornitzky C, Arzey KE, Finlaison D, Hick P, Read A, Hobson-Peters J, May FJ, Doggett SL, Haniotis J, Russell RC, Hall RA, Khromykh AA, Kirkland PD. Characterization of virulent West Nile virus Kunjin strain, Australia, 2011. Emerg Infect Dis 2012 May;18(5):792-800.
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