Archives of Virology

, Volume 156, Issue 2, pp 313–318

The acidic sequence of the NS4A cofactor regulates ATP hydrolysis by the HCV NS3 helicase

  • Sergey A. Shiryaev
  • Andrei V. Chernov
  • Tatiana N. Shiryaeva
  • Alexander E. Aleshin
  • Alex Y. Strongin
Brief Report

Abstract

In flaviviruses and hepatitis C virus (HCV), the NS3 gene encodes the N-terminal protease (NS3pro) and the C-terminal helicase (NS3hel). In HCV, the downstream NS4A is required for the NS3pro activity and exhibits a conserved EFDEMEE motif. To identify the role of this motif, we compared the ATPase and helicase activities of NS3 alone with those of the NS3-NS4A constructs. Our results suggest that the EFDEMEE motif is essential for regulating the ATPase activity of NS3hel. It is likely that this motif interferes with the ATP-binding site of NS3hel. It is becoming clear that NS4A functions as a cofactor of both proteinase and helicase in HCV.

References

  1. 1.
    Dubuisson J (2007) Hepatitis C virus proteins. World J Gastroenterol 13:2406–2415PubMedGoogle Scholar
  2. 2.
    Dumont S, Cheng W, Serebrov V, Beran RK, Tinoco I, Pyle AM, Bustamante C (2006) RNA translocation and unwinding mechanism of HCV NS3 helicase and its coordination by ATP. Nature 439:105–108CrossRefPubMedGoogle Scholar
  3. 3.
    Suzuki R, Suzuki T, Ishii K, Matsuura Y, Miyamura T (1999) Processing and functions of Hepatitis C virus proteins. Intervirology 42:145–152CrossRefPubMedGoogle Scholar
  4. 4.
    Aleshin AE, Shiryaev SA, Strongin AY, Liddington RC (2007) Structural evidence for regulation and specificity of flaviviral proteases and evolution of the Flaviviridae fold. Protein Sci 16:795–806CrossRefPubMedGoogle Scholar
  5. 5.
    Erbel P, Schiering N, D’Arcy A, Renatus M, Kroemer M, Lim SP, Yin Z, Keller TH, Vasudevan SG, Hommel U (2006) Structural basis for the activation of flaviviral NS3 proteases from dengue and West Nile virus. Nat Struct Mol Biol 13:372–373CrossRefPubMedGoogle Scholar
  6. 6.
    Sampath A, Padmanabhan R (2009) Molecular targets for flavivirus drug discovery. Antiviral Res 81:6–15CrossRefPubMedGoogle Scholar
  7. 7.
    Chernov AV, Shiryaev SA, Aleshin AE, Ratnikov BI, Smith JW, Liddington RC, Strongin AY (2008) The two-component NS2B-NS3 proteinase represses DNA unwinding activity of the West Nile virus NS3 helicase. J Biol Chem 283:17270–17278CrossRefPubMedGoogle Scholar
  8. 8.
    Yao N, Reichert P, Taremi SS, Prosise WW, Weber PC (1999) Molecular views of viral polyprotein processing revealed by the crystal structure of the hepatitis C virus bifunctional protease–helicase. Structure 7:1353–1363CrossRefPubMedGoogle Scholar
  9. 9.
    Beran RK, Pyle AM (2008) Hepatitis C viral NS3-4A protease activity is enhanced by the NS3 helicase. J Biol Chem 283:29929–29937CrossRefPubMedGoogle Scholar
  10. 10.
    Shiryaev SA, Chernov AV, Aleshin AE, Shiryaeva TN, Strongin AY (2009) NS4A regulates the ATPase activity of the NS3 helicase: a novel cofactor role of the non-structural protein NS4A from West Nile virus. J Gen Virol 90:2081–2085CrossRefPubMedGoogle Scholar
  11. 11.
    Beran RK, Lindenbach BD, Pyle AM (2009) The NS4A protein of hepatitis C virus promotes RNA-coupled ATP hydrolysis by the NS3 helicase. J Virol 83:3268–3275CrossRefPubMedGoogle Scholar
  12. 12.
    Sikora B, Chen Y, Lichti CF, Harrison MK, Jennings TA, Tang Y, Tackett AJ, Jordan JB, Sakon J, Cameron CE, Raney KD (2008) Hepatitis C virus NS3 helicase forms oligomeric structures that exhibit optimal DNA unwinding activity in vitro. J Biol Chem 283:11516–11525CrossRefPubMedGoogle Scholar
  13. 13.
    Tackett AJ, Chen Y, Cameron CE, Raney KD (2005) Multiple full-length NS3 molecules are required for optimal unwinding of oligonucleotide DNA in vitro. J Biol Chem 280:10797–10806CrossRefPubMedGoogle Scholar
  14. 14.
    Mackintosh SG, Lu JZ, Jordan JB, Harrison MK, Sikora B, Sharma SD, Cameron CE, Raney KD, Sakon J (2006) Structural and biological identification of residues on the surface of NS3 helicase required for optimal replication of the hepatitis C virus. J Biol Chem 281:3528–3535CrossRefPubMedGoogle Scholar
  15. 15.
    Xu T, Sampath A, Chao A, Wen D, Nanao M, Chene P, Vasudevan SG, Lescar J (2005) Structure of the Dengue virus helicase/nucleoside triphosphatase catalytic domain at a resolution of 2.4 A. J Virol 79:10278–10288CrossRefPubMedGoogle Scholar
  16. 16.
    Luo D, Xu T, Hunke C, Gruber G, Vasudevan SG, Lescar J (2008) Crystal structure of the NS3 protease–helicase from dengue virus. J Virol 82:173–183CrossRefPubMedGoogle Scholar
  17. 17.
    Assenberg R, Mastrangelo E, Walter TS, Verma A, Milani M, Owens RJ, Stuart DI, Grimes JM, Mancini EJ (2009) Crystal structure of a novel conformational state of the flavivirus NS3 protein: implications for polyprotein processing and viral replication. J Virol 83:12895–12906CrossRefPubMedGoogle Scholar
  18. 18.
    Roosendaal J, Westaway EG, Khromykh A, Mackenzie JM (2006) Regulated cleavages at the West Nile virus NS4A-2K-NS4B junctions play a major role in rearranging cytoplasmic membranes and Golgi trafficking of the NS4A protein. J Virol 80:4623–4632CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Sergey A. Shiryaev
    • 1
  • Andrei V. Chernov
    • 1
  • Tatiana N. Shiryaeva
    • 1
  • Alexander E. Aleshin
    • 1
  • Alex Y. Strongin
    • 1
  1. 1.Inflammatory and Infectious Disease CenterSanford-Burnham Medical Research InstituteLa JollaUSA

Personalised recommendations