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PDBsum entry 2giq
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References listed in PDB file
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Key reference
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Title
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Selection and characterization of replicon variants dually resistant to thumb- And palm-Binding nonnucleoside polymerase inhibitors of the hepatitis c virus.
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Authors
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S.Le pogam,
H.Kang,
S.F.Harris,
V.Leveque,
A.M.Giannetti,
S.Ali,
W.R.Jiang,
S.Rajyaguru,
G.Tavares,
C.Oshiro,
T.Hendricks,
K.Klumpp,
J.Symons,
M.F.Browner,
N.Cammack,
I.Nájera.
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Ref.
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J Virol, 2006,
80,
6146-6154.
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PubMed id
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Abstract
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Multiple nonnucleoside inhibitor binding sites have been identified within the
hepatitis C virus (HCV) polymerase, including in the palm and thumb domains.
After a single treatment with a thumb site inhibitor (thiophene-2-carboxylic
acid NNI-1), resistant HCV replicon variants emerged that contained mutations at
residues Leu419, Met423, and Ile482 in the polymerase thumb domain. Binding
studies using wild-type (WT) and mutant enzymes and structure-based modeling
showed that the mechanism of resistance is through the reduced binding of the
inhibitor to the mutant enzymes. Combined treatment with a thumb- and a
palm-binding polymerase inhibitor had a dramatic impact on the number of
replicon colonies able to replicate in the presence of both inhibitors. A more
exact characterization through molecular cloning showed that 97.7% of replicons
contained amino acid substitutions that conferred resistance to either of the
inhibitors. Of those, 65% contained simultaneously multiple amino acid
substitutions that conferred resistance to both inhibitors. Double-mutant
replicons Met414Leu and Met423Thr were predominantly selected, which showed
reduced replication capacity compared to the WT replicon. These findings
demonstrate the selection of replicon variants dually resistant to two NS5B
polymerase inhibitors binding to different sites of the enzyme. Additionally,
these findings provide initial insights into the in vitro mutational threshold
of the HCV NS5B polymerase and the potential impact of viral fitness on the
selection of multiple-resistant mutants.
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