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Identification of Peptide Leads to Inhibit Hepatitis C Virus: Inhibitory Effect of Plectasin Peptide Against Hepatitis C Serine Protease

  • Ammar Y. Abdulrahman
  • Hussin A. Rothan
  • Nurshamimi Nor Rashid
  • See Khai Lim
  • Wajihah Sakhor
  • Kah Ching Tee
  • Teow Chong Teoh
  • Noorsaadah A. Rahman
  • Rohana Yusof
Article

Abstract

The emerging of hepatitis C virus (HCV) resistant strains has been considered as a main drawback of the available drugs. Since HCV has a large inactive surface, we would like to hypothesis that the mutation occur in HCV is minimal and causing less resistance against inhibition. In this study, a short peptide inhibitor of HCV namely plectasin was identified by HCV NS3-4A serine protease assay. Plectasin peptide showed considerable inhibition against HCV NS3-4A serine protease. Enzymatic activity of the recombinant NS3-4Apro was analysed by fluorescence release from several fluorogenic peptide substrates which resembling the dibasic cleavage site sequences of the flavivirus polyprotein precursor. Of all amc-labelled peptides, Pyr-RTKR-amc was the most efficiently cleaved substrate with the lowest Km value of 20 µM. The kinetic assay showed that plectasin peptide inhibited NS3-4Apro activity with an IC50 value of 4.3 μM compared to the aprotinin as a standard proteases inhibitor with an IC50 of 6.1 μM. From the results, plectasin peptide also demonstrated a dose-dependent inhibition of HCV replication with a considerable reduction in RLuc activity at 15 µM using HCV replicon- containing Huh-7 cells. Our study has identified a unique natural peptide that can be used to highlight novel structures for the development of drug derivatives with high efficacy of HCV NS3-4A protease inhibitors.

Keywords

Antiviral peptide Plectasin Hepatitis C virus NS3-4A serine protease HCV replicon assay 

Notes

Acknowledgments

This study was sponsored by the Ministry of Higher education/Malaysia through the University of Malaya, TRGS Grant (TR0001D-2014B).

Compliance with Ethical Standards

Conflict of Interest

Authors declare that they have no conflict of interest.

Ethical Approval

This article does not contain any study with human or animals performed by any of the authors.

Supplementary material

10989_2016_9544_MOESM1_ESM.docx (14 kb)
Supplementary material 1 (DOCX 14 kb)

References

  1. Alexopoulou A, Papatheodoridis GV (2012) Current progress in the treatment of chronic hepatitis C. World J Gastroenterol 18(42):6060–6069. doi: 10.3748/wjg.v18.i42.6060 CrossRefPubMedPubMedCentralGoogle Scholar
  2. Ashfaq UA, Javed T, Rehman S, Nawaz Z, Riazuddin S (2011) An overview of HCV molecular biology, 21 replication, and immune responses. Virol J 8. doi: 10.1186/1743-422x-8-161Google Scholar
  3. Asselah T (2014) Daclatasvir plus sofosbuvir for HCV infection: an oral combination therapy with high antiviral efficacy. J Hepatol 61(2):435–438. doi: 10.1016/j.jhep.2014.04.042 CrossRefPubMedGoogle Scholar
  4. Bhatia HK, Singh H, Grewal N, Natt NK (2014) Sofosbuvir: a novel treatment option for chronic hepatitis C infection. J Pharmacol Pharmacother 5(4):278–284. doi: 10.4103/0976-500X.142464 CrossRefPubMedPubMedCentralGoogle Scholar
  5. Brinch KS, Frimodt-Moller N, Hoiby N, Kristensen HH (2009) Influence of antidrug antibodies on plectasin efficacy and pharmacokinetics. Antimicrob Agents Chemother 53(11):4794–4800. doi: 10.1128/AAC.00440-09 CrossRefPubMedPubMedCentralGoogle Scholar
  6. Caillet-Saguy C, Lim SP, Shi PY, Lescar J, Bressanelli S (2014) Polymerases of hepatitis C viruses and flaviviruses: structural and mechanistic insights and drug development. Antiviral Res 105:8–16. doi: 10.1016/j.antiviral.2014.02.006 CrossRefPubMedGoogle Scholar
  7. Carlson A, Gregorich Z, Striker R (2013) Telaprevir to boceprevir switch highlights lack of cross-reactivity. Clin Infect Dis 56(4):552–554. doi: 10.1093/cid/cis960 CrossRefPubMedGoogle Scholar
  8. de Vries SJ, van Dijk M, Bonvin AM (2010) The HADDOCK web server for data-driven biomolecular docking. Nat Protoc 5(5):883–897. doi: 10.1038/nprot.2010.32 CrossRefPubMedGoogle Scholar
  9. Di Marco S et al (2000) Inhibition of the hepatitis C virus NS3/4A protease. The crystal structures of two protease-inhibitor complexes. J Biol Chem 275(10):7152–7157CrossRefPubMedGoogle Scholar
  10. Gower E, Estes C, Blach S, Razavi-Shearer K, Razavi H (2014) Global epidemiology and genotype distribution of the hepatitis C virus infection. J Hepatol 61(1 Suppl):S45–S57. doi: 10.1016/j.jhep.2014.07.027 CrossRefPubMedGoogle Scholar
  11. Gozdek A et al (2008) NS3 Peptide, a novel potent hepatitis C virus NS3 helicase inhibitor: its mechanism of action and antiviral activity in the replicon system. Antimicrob Agents Chemother 52(2):393–401. doi: 10.1128/AAC.00961-07 CrossRefPubMedGoogle Scholar
  12. Krieger E, Koraimann G, Vriend G (2002) Increasing the precision of comparative models with YASARA NOVA–a self-parameterizing force field. Proteins 47(3):393–402CrossRefPubMedGoogle Scholar
  13. Kugler J et al (2012) High affinity peptide inhibitors of the hepatitis C virus NS3-4A protease refractory to common resistant mutants. J Biol Chem 287(46):39224–39232. doi: 10.1074/jbc.M112.393843 CrossRefPubMedPubMedCentralGoogle Scholar
  14. Lavanchy D (2011) Evolving epidemiology of hepatitis C virus. Clin Microbiol Infect 17(2):107–115. doi: 10.1111/j.1469-0691.2010.03432.x CrossRefPubMedGoogle Scholar
  15. Liang QL, Zhou K, He HX (2010) Retrocyclin 2: a new therapy against avian influenza H5N1 virus in vivo and vitro. Biotechnol Lett 32(3):387–392. doi: 10.1007/s10529-009-0167-2 CrossRefPubMedGoogle Scholar
  16. Mandal K et al (2009) Racemic crystallography of synthetic protein enantiomers used to determine the X-ray structure of plectasin by direct methods. Protein Sci 18(6):1146–1154. doi: 10.1002/pro.127 CrossRefPubMedPubMedCentralGoogle Scholar
  17. Marks KM, Jacobson IM (2012) The first wave: HCV NS3 protease inhibitors telaprevir and boceprevir. Antivir Ther 17(6):1119–1131. doi: 10.3851/IMP2424 CrossRefPubMedGoogle Scholar
  18. Mayhoub AS (2012) Hepatitis C RNA-dependent RNA polymerase inhibitors: a review of structure-activity and resistance relationships; different scaffolds and mutations. Bioorg Med Chem 20(10):3150–3161. doi: 10.1016/j.bmc.2012.03.049 CrossRefPubMedGoogle Scholar
  19. Morikawa K et al (2011) Nonstructural protein 3-4A: the Swiss army knife of hepatitis C virus. J Viral Hepat 18(5):305–315. doi: 10.1111/j.1365-2893.2011.01451.x CrossRefPubMedGoogle Scholar
  20. Mygind PH et al (2005) Plectasin is a peptide antibiotic with therapeutic potential from a saprophytic fungus. Nature 437(7061):975–980. doi: 10.1038/nature04051 CrossRefPubMedGoogle Scholar
  21. Petta S et al (2014) Cost-effectiveness of sofosbuvir-based triple therapy for untreated patients with genotype 1 chronic hepatitis C. Hepatology 59(5):1692–1705. doi: 10.1002/hep.27010 CrossRefPubMedGoogle Scholar
  22. Raney KD, Sharma SD, Moustafa IM, Cameron CE (2010) Hepatitis C virus non-structural protein 3 (HCV NS3): a multifunctional antiviral target. J Biol Chem 285(30):22725–22731. doi: 10.1074/jbc.R110.125294 CrossRefPubMedPubMedCentralGoogle Scholar
  23. Romano KP et al (2012) The molecular basis of drug resistance against hepatitis C virus NS3/4A protease inhibitors. PLoS Pathog 8(7):e1002832. doi: 10.1371/journal.ppat.1002832 CrossRefPubMedPubMedCentralGoogle Scholar
  24. Rosenquist A et al (2014) Discovery and development of simeprevir (TMC435), a HCV NS3/4A protease inhibitor. J Med Chem 57(5):1673–1693. doi: 10.1021/jm401507s CrossRefPubMedGoogle Scholar
  25. Rothan HA, Abdulrahman AY, Sasikumer PG, Othman S, Rahman NA, Yusof R (2012a) Protegrin-1 inhibits dengue NS2B-NS3 serine protease and viral replication in MK2 cells. J Biomed Biotechnol 2012:251482. doi: 10.1155/2012/251482 CrossRefPubMedPubMedCentralGoogle Scholar
  26. Rothan HA, Han HC, Ramasamy TS, Othman S, Rahman NA, Yusof R (2012b) Inhibition of dengue NS2B-NS3 protease and viral replication in Vero cells by recombinant retrocyclin-1. BMC Infect Dis 12:314. doi: 10.1186/1471-2334-12-314 CrossRefPubMedPubMedCentralGoogle Scholar
  27. Schneider MD, Sarrazin C (2014) Antiviral therapy of hepatitis C in 2014: do we need resistance testing? Antivir Res 105:64–71. doi: 10.1016/j.antiviral.2014.02.011 CrossRefPubMedGoogle Scholar
  28. Simmonds P (2004) Genetic diversity and evolution of hepatitis C virus–15 years on. J Gen Virol 85(Pt 11):3173–3188. doi: 10.1099/vir.0.80401-0 CrossRefPubMedGoogle Scholar
  29. Soumana DI, Ali A, Schiffer CA (2014) Structural analysis of asunaprevir resistance in HCV NS3/4A protease. ACS Chem Biol 9(11):2485–2490. doi: 10.1021/cb5006118 CrossRefPubMedPubMedCentralGoogle Scholar
  30. Thorlund K, Druyts E, El Khoury AC, Mills EJ (2012) Budget impact analysis of boceprevir and telaprevir for the treatment of hepatitis C genotype 1 infection. Clinicoecon Outcomes Res 4:349–359. doi: 10.2147/CEOR.S37205 PubMedPubMedCentralGoogle Scholar
  31. Uhlig T et al (2014) The emergence of peptides in the pharmaceutical business: from exploration to exploitation. EuPA Open Proteom 4:58–69. doi: 10.1016/j.euprot.2014.05.003 CrossRefGoogle Scholar
  32. Wassenaar TA et al (2012) WeNMR: structural biology on the grid. J Grid Comput 10(4):743–767. doi: 10.1007/s10723-012-9246-z CrossRefGoogle Scholar
  33. Yu Y et al (2014) Discovering novel anti-HCV compounds with inhibitory activities toward HCV NS3/4A protease. Acta Pharmacol Sin 35(8):1074–1081. doi: 10.1038/aps.2014.55 CrossRefPubMedPubMedCentralGoogle Scholar
  34. Zhang J, Yang Y, Teng D, Tian Z, Wang S, Wang J (2011) Expression of plectasin in Pichia pastoris and its characterization as a new antimicrobial peptide against Staphyloccocus and Streptococcus. Protein Expr Purif 78(2):189–196. doi: 10.1016/j.pep.2011.04.014 CrossRefPubMedGoogle Scholar
  35. Zhang Q et al (2012) Activation of the Ras/Raf/MEK pathway facilitates hepatitis C virus replication via attenuation of the interferon-JAK-STAT pathway. J Virol 86(3):1544–1554. doi: 10.1128/JVI.00688-11 CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Ammar Y. Abdulrahman
    • 1
  • Hussin A. Rothan
    • 1
  • Nurshamimi Nor Rashid
    • 1
  • See Khai Lim
    • 3
  • Wajihah Sakhor
    • 1
  • Kah Ching Tee
    • 1
  • Teow Chong Teoh
    • 2
  • Noorsaadah A. Rahman
    • 4
  • Rohana Yusof
    • 1
  1. 1.Department of Molecular Medicine, Faculty of MedicineUniversity of MalayaKuala LumpurMalaysia
  2. 2.Faculty of Science, Institute of Biological SciencesUniversity of MalayaKuala LumpurMalaysia
  3. 3.Department of Pharmacy, Faculty of MedicineUniversity of MalayaKuala LumpurMalaysia
  4. 4.Department of Chemistry, Faculty of ScienceUniversity of MalayaKuala LumpurMalaysia

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