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Biotechnology Letters

, Volume 40, Issue 2, pp 315–324 | Cite as

Development of RNA aptamer that inhibits methyltransferase activity of dengue virus

Original Research Paper

Abstract

Objectives

To develop an RNA aptamer specific for the methyltransferase (MTase) of dengue virus (DENV) which is essential for viral genome replication and translation acting directly on N-7 and 2′-O-methylation of the type-I cap structure of the viral RNA.

Results

We identified 2′-fluoro-modified RNA aptamers that can specifically bind DENV serotype 2 (DENV2) MTase using systematic evolution of ligands by exponential enrichment technology. We truncated the chosen aptamer into a 45-mer RNA sequence that can bind DENV2 MTase with K d  ~ 28 nM and inhibit N-7 methylation activity of the protein. Moreover, the 45-mer truncated aptamer could not only bind with an K d  ~ 15.6 nM but also inhibit methylation activity of DENV serotype 3 (DENV3) MTase. The 45-mer aptamer competitively impeded binding of both DENV2 and DENV3 genomic RNA to MTase of each serotype.

Conclusion

The selected 45-mer truncated RNA aptamer specifically and avidly bound DENV MTase and competitively inhibited its methylation activity, and thus could be useful for the development of anti-DENV agents.

Keywords

Antiviral agent Dengue virus Methyltransferase RNA aptamer 

Notes

Acknowledgements

The present research was conducted by the research fund of Dankook University in Republic of Korea in 2015.

Supporting information

Supplementary Table 1—Sequence of primers.

Supplementary Fig. 1—Enrichment of selected RNA aptamer pool.

Supplementary Fig. 2—Aptamer truncation experiment.

Supplementary Fig. 3—Comparison of amino acid sequence of MTase protein of DENV 1-4 serotypes.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflicts of interest.

Supplementary material

10529_2017_2462_MOESM1_ESM.pdf (821 kb)
Supplementary material 1 (PDF 820 kb)

References

  1. Coleman TM, Wang G, Huang F (2004) Superior 5′ homogeneity of RNA from ATP-initiated transcription under the T7 φ2.5 promoter. Nucleic Acids Res 32:e14CrossRefPubMedPubMedCentralGoogle Scholar
  2. Daffis S et al (2010) 2′-O methylation of the viral mRNA cap evades host restriction by IFIT family members. Nature 468:452–456CrossRefPubMedPubMedCentralGoogle Scholar
  3. Diamond MS, Pierson TC (2015) Molecular insight into dengue virus pathogenesis and its implications for disease control. Cell 162:488–492CrossRefPubMedPubMedCentralGoogle Scholar
  4. Dong H, Ray D, Ren S, Zhang B, Puig-Basagoiti F, Takagi Y, Ho CK, Li H, Shi PY (2007) Distinct RNA elements confer specificity to flavivirus RNA cap methylation events. J Virol 81:4412–4421CrossRefPubMedPubMedCentralGoogle Scholar
  5. Dong H, Zhang B, Shi PY (2008) Flavivirus methyltransferase: a novel antiviral target. Antivir Res 80:1–10CrossRefPubMedPubMedCentralGoogle Scholar
  6. Dong H, Chang DC, Xie X, Toh YX, Chung KY, Zou G, Lescar J, Lim SP, Shi PY (2010) Biochemical and genetic characterization of dengue virus methyltransferase. Virology 405:568–578CrossRefPubMedGoogle Scholar
  7. Durbin AP (2016) A dengue vaccine. Cell 166:1CrossRefPubMedGoogle Scholar
  8. García-Sastre A (2011) 2 Methylate or not 2 methylate: viral evasion of the type I interferon response. Nat Immunol 2:114–115CrossRefGoogle Scholar
  9. Han SR, Lee SW (2017) Inhibition of japanese encephalitis virus (JEV) replication by specific RNA aptamer against JEV methyltransferase. Biochem Biophys Res Commun 483:687–693CrossRefPubMedGoogle Scholar
  10. Iglesias NG, Filomatori CV, Gamarnik AV (2011) The F1 motif of dengue virus polymerase NS5 is involved in promoter-dependent RNA synthesis. J Virol 85:5745–5756CrossRefPubMedPubMedCentralGoogle Scholar
  11. Jung SN, Kang SK, Yeo GH, Li HY, Jiang T, Nah JW, Bok JD, Cho CS, Choi YJ (2015) Targeted delivery of vaccine to dendritic cells by chitosan nanoparticles conjugated with a targeting peptide ligand selected by phage display technique. Macromol Biosci 15:395–404CrossRefPubMedGoogle Scholar
  12. Keefe AD, Pai S, Ellington A (2010) Aptamers as therapeutics. Nat Rev Drug Discov 9:537–550CrossRefPubMedGoogle Scholar
  13. Kim SH, Yang IY, Jang SH, Kim J, Truong TT, Van Pham T, Truong NU, Lee KY, Jang YS (2013) C5a receptor-targeting ligand-mediated delivery of dengue virus antigen to M cells evokes antigen-specific systemic and mucosal immune responses in oral immunization. Microbes Infect 15:895–902CrossRefPubMedGoogle Scholar
  14. Kroschewski H (2008) Mutagenesis of the dengue virus type 2 NS5 methyltransferase domain. J Biol Chem 283:19410–19421CrossRefPubMedGoogle Scholar
  15. Lee CH, Lee YJ, Kim JH, Lim JH, Kim JH, Han W, Lee SH, Noh GJ, Lee SW (2013) Inhibition of hepatitis C virus (HCV) replication by specific RNA aptamers against HCV NS5B RNA replicase. J Virol 87:7064–7074CrossRefPubMedPubMedCentralGoogle Scholar
  16. Mukhopadhyay S, Kuhn RJ, Rossmann MG (2005) A structural perspective of the flavivirus life cycle. Nat Rev Microbiol 3:13–22CrossRefPubMedGoogle Scholar
  17. Potisopon S et al (2014) The methyltransferase domain of dengue virus protein NS5 ensures efficient RNA synthesis initiation and elongation by the polymerase domain. Nucleic Acids Res 42:11642–11656CrossRefPubMedPubMedCentralGoogle Scholar
  18. Tuerk C, Gold L (1990) Systematic evolution of ligands by exponential enrichment: RNA ligands to bacteriophage T4 DNA polymerase. Science 249:505–510CrossRefPubMedGoogle Scholar
  19. Whitehead SS, Blaney JE, Durbin AP, Murphy BR (2007) Prospects for a dengue virus vaccine. Nat Rev Microbiol 5:518–528CrossRefPubMedGoogle Scholar
  20. Zhou Y, Ray D, Zhao Y, Dong H, Ren S, Li Z, Guo Y, Bernard KA, Shi PY, Li H (2007) Structure and function of flavivirus NS5 methyltransferase. J Virol 81:3891–3903CrossRefPubMedPubMedCentralGoogle Scholar
  21. Zuker M (2003) Mfold web server for nucleic acid folding and hybridization prediction. Nucleic Acids Res 31:3406–3415CrossRefPubMedPubMedCentralGoogle Scholar
  22. Züst R et al (2011) Ribose 2′-O-methylation provides a molecular signature for the distinction of self and non-self mRNA dependent on the RNA sensor Mda5. Nat Immunol 12:137–143CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2017

Authors and Affiliations

  1. 1.Department of Integrated Life Sciences, Research Institute of Advanced OmicsDankook UniversityYonginRepublic of Korea

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