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Title
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Crystal Structures of Clinically Relevant Lys103Asn/Tyr181Cys Double Mutant HIV-1 Reverse Transcriptase in Complexes with ATP and Non-nucleoside Inhibitor HBY 097.
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Authors
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K.Das,
S.G.Sarafianos,
A.D.Clark,
P.L.Boyer,
S.H.Hughes,
E.Arnold.
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Ref.
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J Mol Biol, 2007,
365,
77-89.
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PubMed id
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Abstract
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Lys103Asn and Tyr181Cys are the two mutations frequently observed in patients
exposed to various non-nucleoside reverse transcriptase inhibitor drugs
(NNRTIs). Human immunodeficiency virus (HIV) strains containing both reverse
transcriptase (RT) mutations are resistant to all of the approved NNRTI drugs.
We have determined crystal structures of Lys103Asn/Tyr181Cys mutant HIV-1 RT
with and without a bound non-nucleoside inhibitor (HBY 097,
(S)-4-isopropoxycarbonyl-6-methoxy-3-(methylthio-methyl)-3,4-dihydroquinoxalin-2(1H)-thione)
at 3.0 A and 2.5 A resolution, respectively. The structure of the double mutant
RT/HBY 097 complex shows a rearrangement of the isopropoxycarbonyl group of HBY
097 compared to its binding with wild-type RT. HBY 097 makes a hydrogen bond
with the thiol group of Cys181 that helps the drug retain potency against the
Tyr181Cys mutation. The structure of the unliganded double mutant HIV-1 RT
showed that Lys103Asn mutation facilitates coordination of a sodium ion with
Lys101 O, Asn103 N and O(delta1), Tyr188 O(eta), and two water molecules. The
formation of the binding pocket requires the removal of the sodium ion. Although
the RT alone and the RT/HBY 097 complex were crystallized in the presence of
ATP, only the RT has an ATP coordinated with two Mn(2+) at the polymerase active
site. The metal coordination mimics a reaction intermediate state in which
complete octahedral coordination was observed for both metal ions. Asp186
coordinates at an axial position whereas the carboxylates of Asp110 and Asp185
are in the planes of coordination of both metal ions. The structures provide
evidence that NNRTIs restrict the flexibility of the YMDD loop and prevent the
catalytic aspartate residues from adopting their metal-binding conformations.
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