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Mutations that affect dimer formation and helicase activity of the hepatitis C virus helicase

Academic Article
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Overview

authors

  • Khu, Y. L.
  • Koh, E.
  • Lim, S. P.
  • Tan, Y. H.
  • Brenner, Sydney
  • Lim, S. G.
  • Hong, W. J.
  • Goh, P. Y.

publication date

  • 2001

journal

  • Journal of Virology  Journal

abstract

  • Interaction between viral proteins is necessary for viral replication and viral particle assembly. We used the yeast two-hybrid assay to identify interactions among all the mature proteins of the hepatitis C virus. The interaction between NS3 and NS3 was one of the strongest viral protein-protein interactions detected. The minimal region required for this interaction was mapped to a specific subdomain of 174 amino acids in the N terminus of the helicase region. Random mutations in the minimal region were generated by PCR, and mutants that failed to interact with a wild-type minimal fragment were isolated using the yeast two-hybrid assay as a screen. Three of these mutations resulted in a reduction or a loss of interaction between helicases. Analytical gel filtration showed that in the presence of an oligonucleotide, wild-type helicases form dimers whereas the mutants remain mostly monomeric. All three mutants were partially or almost inactive when assayed for helicase activity in vitro. Mixing a mutant helicase (Y267S) with wild-type helicase did not dramatically affect helicase activity. These data indicate that dimerization of the helicase is important for helicase activity. The mutations that reduce self-association of the helicase may define the key residues involved in NS3-NS3 dimerization.

subject areas

  • Amino Acid Sequence
  • Animals
  • COS Cells
  • Dimerization
  • Molecular Sequence Data
  • Mutation
  • RNA Helicases
  • Viral Nonstructural Proteins
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Identity

PubMed Central ID

  • PMC113914

International Standard Serial Number (ISSN)

  • 0022-538X

Digital Object Identifier (DOI)

  • 10.1128/jvi.75.1.205-214.2001

PubMed ID

  • 11119590
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Additional Document Info

start page

  • 205

end page

  • 214

volume

  • 75

issue

  • 1

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