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* Residue conservation analysis
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References listed in PDB file
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Key reference
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Title
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Crystal structures of 8-Cl and 9-Cl tibo complexed with wild-Type HIV-1 rt and 8-Cl tibo complexed with the tyr181cys HIV-1 rt drug-Resistant mutant.
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Authors
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K.Das,
J.Ding,
Y.Hsiou,
A.D.Clark,
H.Moereels,
L.Koymans,
K.Andries,
R.Pauwels,
P.A.Janssen,
P.L.Boyer,
P.Clark,
R.H.Smith,
M.B.Kroeger smith,
C.J.Michejda,
S.H.Hughes,
E.Arnold.
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Ref.
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J Mol Biol, 1996,
264,
1085-1100.
[DOI no: ]
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PubMed id
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Abstract
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Human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) is an
important target for chemotherapeutic agents used in the treatment of AIDS; the
TIBO compounds are potent non-nucleoside inhibitors of HIV-1 RT (NNRTIs).
Crystal structures of HIV-1 RT complexed with 8-Cl TIBO (R86183, IC50 = 4.6 nM)
and 9-Cl TIBO (R82913, IC50 = 33 nM) have been determined at 3.0 A resolution.
Mutant HIV-1 RT, containing Cys in place of Tyr at position 181 (Tyrl81Cys), is
highly resistant to many NNRTIs and HIV-1 variants containing this mutation have
been selected in both cell culture and clinical trials. We also report the
crystal structure of Tyrl81Cys HIV-1 RT in complex with 8-Cl TIBO (IC50 = 130
nM) determined at 3.2 A resolution. Averaging of the electron density maps
computed for different HIV-1 RT/NNRTI complexes and from diffraction datasets
obtained using a synchrotron source from frozen (-165 degrees C) and cooled (-10
degrees C) crystals of the same complex was employed to improve the quality of
electron density maps and to reduce model bias. The overall locations and
conformations of the bound inhibitors in the complexes containing wild-type
HIV-1 RT and the two TIBO inhibitors are very similar, as are the overall shapes
and volumes of the non-nucleoside inhibitor-binding pocket (NNIBP). The major
differences between the two wild-type HIV-1 RT/TIBO complexes occur in the
vicinity of the TIBO chlorine substituents and involve the polypeptide segments
around the beta5-beta6 connecting loop (residues 95 to 105) and the
beta13-beta14 hairpin (residues 235 and 236). In all known structures of HIV-1
RT/NNRTI complexes, including these two, the position of the beta12-beta13
hairpin or the "primer grip" is significantly displaced relative to the position
in the structure of HIV-1 RT complexed with a double-stranded DNA and in
unliganded HIV-1 RT structures. Since the primer grip helps to position the
template-primer, this displacement suggests that binding of NNRTIs would affect
the relative positions of the primer terminus and the polymerase active site.
This could explain biochemical data showing that NNRTI binding to HIV-1 RT
reduces efficiency of the chemical step of DNA polymerization, but does not
prevent binding of either dNTPs or DNA. When the structure of the Tyr181Cys
mutant HIV-1 RT in complex with 8-Cl TIBO is compared with the corresponding
structure containing wild-type HIV-1 RT, the overall conformations of Tyr181Cys
and wild-type HIV-1 RT and of the 8-Cl TIBO inhibitors are very similar. Some
positional changes in the polypeptide backbone of the beta6-beta10-beta9 sheet
containing residue 181 are observed when the Tyr181Cys and wild-type complexes
are compared, particularlty near residue Val179 of beta9. In the p51 subunit,
the Cys181 side-chain is oriented in a similar direction to the Tyr181
side-chain in the wild-type complex. However, the electron density corresponding
to the sulfur of the Cys181 side-chain in the p66 subunit is very weak,
indicating that the thiol group is disordered, presumably because there is no
significant interaction with either 8-Cl TIBO or nearby amino acid residues. In
the mutant complex, there are slight rearrangements of the side-chains of other
amino acid residues in the NNIBP and of the flexible dimethylallyl group of 8-Cl
TIBO; these conformational changes could potentially compensate for the
interactions that were lost when the relatively large tyrosine at position 181
was replaced by a less bulky cysteine residue. In the corresponding wild-type
complex, Tyr181 iin the p66 subunit has significant interactions with the bound
inhibitor and the position of the Tyr181 side-chain is well defined in both
subunits. Apparently the Tyr181 --> Cys mutation eliminates favorable
contacts of the aromatic ring of the tyrosine and the bou
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Figure 1.
Figure 1. Chemical structures
with the numbering scheme used
and distances (E3.6 Å ) between
atoms of the TIBO inhibitor and of
the amino acid residues of the
NNIBP for: (a) 8-Cl TIBO (R86183,
tivirapine) complexed with wild-
type HIV-1 RT; (b) 8-Cl TIBO
complexed with Tyr181Cys mutant
HIV-1 RT; and (c) 9-Cl TIBO
(R82913) complexed with wild-type
HIV-1 RT. An NNIBP residue is
shown only if atoms of that residue
are E3.6 Å from an inhibitor atom
with the exception of Cys181 in (b).
The wings I and II portions of the
inhibitors in the butterfly-like anal-
ogy for NNRTIs (Ding et al., 1995a)
are indicated here and in sub-
sequent Figures by Roman nu-
merals I and II. The dotted line in (a)
indicates the subdivision of atoms
between wings I and II.
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Figure 5.
Figure 5. A stereoview of the superposition (based on the C
a
atoms of the b6-b10-b9 sheet) of the HIV-1 RT/DNA/Fab
complex structure (in gray) (Jacobo-Molina et al., 1993) on the HIV-1 RT/9-Cl TIBO complex structure (in cyan) in the
regions near the NNIBP and the polymerase active site showing the disposition of the b12-b13-b14 sheet containing
the primer grip. Bound 9-Cl TIBO in the HIV-1 RT/9-Cl TIBO complex is shown in gold and the two 3'-terminal
nucleotides 17 and 18 of the primer strand in the HIV-1 RT/DNA/Fab complex are shown with a yellow ball-and-stick
model. The broken line represents interactions between the primer grip and the primer terminal phosphate in the HIV-1
RT/DNA/Fab complex and the arrow indicates the movement of the primer grip that accompanies NNRTI binding.
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The above figures are
reprinted
by permission from Elsevier:
J Mol Biol
(1996,
264,
1085-1100)
copyright 1996.
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Secondary reference #1
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Title
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Targeting HIV reverse transcriptase for anti-Aids drug design: structural and biological considerations for chemotherapeutic strategies.
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Authors
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E.Arnold,
K.Das,
J.Ding,
P.N.Yadav,
Y.Hsiou,
P.L.Boyer,
S.H.Hughes.
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Ref.
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Drug Des Discov, 1996,
13,
29-47.
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PubMed id
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Secondary reference #2
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Title
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Structure of unliganded HIV-1 reverse transcriptase at 2.7 a resolution: implications of conformational changes for polymerization and inhibition mechanisms.
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Authors
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Y.Hsiou,
J.Ding,
K.Das,
A.D.Clark,
S.H.Hughes,
E.Arnold.
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Ref.
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Structure, 1996,
4,
853-860.
[DOI no: ]
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PubMed id
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Figure 3.
Figure 3. Superposition of (a) unliganded RT and RT–DNA–Fab
complex and (b) unliganded RT and RT–α-APA
(α-anilinophenylacetamide) complex based on 89 Cα atoms in the
p66 palm subdomain, including the β6–β10–β9 region. The
unliganded RT is shown in red, RT–α-APA in blue, and
RT–DNA–Fab in green. A comparison of the two superpositions
reveals that NNRTI binding appears to be accompanied by a
long-range distortion that is coupled with a hinge motion
(indicated by curved arrows) between the β6–β10–β9 and
β12–β13–β14 sheets at the p66 palm subdomain (within the
circle). The different positions of the thumb in different HIV-1
RT structures supports the idea that this subdomain could play
a role during polymerization. Figure 3. Superposition of (a)
unliganded RT and RT–DNA–Fab complex and (b) unliganded RT
and RT–α-APA (α-anilinophenylacetamide) complex based on 89
Cα atoms in the p66 palm subdomain, including the
β6–β10–β9 region. The unliganded RT is shown in red,
RT–α-APA in blue, and RT–DNA–Fab in green. A comparison
of the two superpositions reveals that NNRTI binding appears to
be accompanied by a long-range distortion that is coupled with a
hinge motion (indicated by curved arrows) between the
β6–β10–β9 and β12–β13–β14 sheets at the p66 palm
subdomain (within the circle). The different positions of the
thumb in different HIV-1 RT structures supports the idea that
this subdomain could play a role during polymerization.
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Figure 4.
Figure 4. Stereoview of a portion of a (2mF[obs]–F[calc])
difference Fourier map at the p66 connection subdomain, at 2.7
å resolution. The phases were computed from the current
atomic model and the map is contoured at 1.4σ. The side chain
and the carboxyl groups are well defined in the electron density
map. Figure 4. Stereoview of a portion of a
(2mF[obs]–F[calc]) difference Fourier map at the p66
connection subdomain, at 2.7 å resolution. The phases were
computed from the current atomic model and the map is contoured
at 1.4σ. The side chain and the carboxyl groups are well
defined in the electron density map.
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The above figures are
reproduced from the cited reference
with permission from Cell Press
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Secondary reference #3
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Title
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Structure of HIV-1 reverse transcriptase in a complex with the non-Nucleoside inhibitor alpha-Apa r 95845 at 2.8 a resolution.
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Authors
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J.Ding,
K.Das,
C.Tantillo,
W.Zhang,
A.D.Clark,
S.Jessen,
X.Lu,
Y.Hsiou,
A.Jacobo-Molina,
K.Andries.
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Ref.
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Structure, 1995,
3,
365-379.
[DOI no: ]
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PubMed id
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Figure 2.
Figure 2. (a) Stereoview of difference Fourier maps showing the
fit of α-APA into the electron density. The α-APA coordinates
correspond to the current refined model. The green map is the
difference Fourier map at 3.5 å resolution between the
HIV-1 RT/α-APA R 95845 (dibrominated) complex and the HIV-1
RT/α-APA R 90385 (dichlorinated) complex contoured at a 10σ
level which revealed the positions of two bromine atoms. The
blue map is the mF[o]–F[c] difference Fourier map at 3.0
å resolution calculated from a model before α-APA was
included in the refinement; the phases were computed from the
back-transformation of electron density averaged from three
HIV-1 RT/inhibitor complexes; the map is contoured at 1.5σ. (b)
Stereoview of a portion of a 2mF[o]–F[c] difference Fourier
map at 2.8 å resolution in the region around the
non-nucleoside inhibitor-binding pocket showing the α-APA
inhibitor and some of the nearby amino acid residues. The phases
were computed from the current atomic model (R=0.255) and the
map is contoured at 1.2σ. Figure 2. (a) Stereoview of
difference Fourier maps showing the fit of α-APA into the
electron density. The α-APA coordinates correspond to the
current refined model. The green map is the difference Fourier
map at 3.5 å resolution between the HIV-1 RT/α-APA R
95845 (dibrominated) complex and the HIV-1 RT/α-APA R 90385
(dichlorinated) complex contoured at a 10σ level which revealed
the positions of two bromine atoms. The blue map is the
mF[o]–F[c] difference Fourier map at 3.0 å resolution
calculated from a model before α-APA was included in the
refinement; the phases were computed from the
back-transformation of electron density averaged from three
HIV-1 RT/inhibitor complexes; the map is contoured at 1.5σ. (b)
Stereoview of a portion of a 2mF[o]–F[c] difference Fourier
map at 2.8 å resolution in the region around the
non-nucleoside inhibitor-binding pocket showing the α-APA
inhibitor and some of the nearby amino acid residues. The phases
were computed from the current atomic model (R=0.255) and the
map is contoured at 1.2σ.
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Figure 4.
Figure 4. Close-up of the non-nucleoside inhibitor-binding
pocket in the structure of the HIV-1 RT/α-APA complex looking
through a putative entrance to the pocket, showing interactions
between α-APA and nearby amino acid residues. α-APA is shown
in purple as a ball-and-stick model with carbons purple,
nitrogens cyan, oxygens red and bromines magenta. The β7–β8
portion of p51 has a dashed outline. The side chains are shown
for the amino acid residues that make close contacts with α-APA
(in green), and for the three essential aspartic acid residues
D110, D185 and D186 (in red) at the polymerase active site.
Dashed lines indicate connections between the side chains and
the β-strands. Figure 4. Close-up of the non-nucleoside
inhibitor-binding pocket in the structure of the HIV-1 RT/α-APA
complex looking through a putative entrance to the pocket,
showing interactions between α-APA and nearby amino acid
residues. α-APA is shown in purple as a ball-and-stick model
with carbons purple, nitrogens cyan, oxygens red and bromines
magenta. The β7–β8 portion of p51 has a dashed outline. The
side chains are shown for the amino acid residues that make
close contacts with α-APA (in green), and for the three
essential aspartic acid residues D110, D185 and D186 (in red) at
the polymerase active site. Dashed lines indicate connections
between the side chains and the β-strands.
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The above figures are
reproduced from the cited reference
with permission from Cell Press
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Secondary reference #4
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Title
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Structure of HIV-1 rt/tibo r 86183 complex reveals similarity in the binding of diverse nonnucleoside inhibitors.
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Authors
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J.Ding,
K.Das,
H.Moereels,
L.Koymans,
K.Andries,
P.A.Janssen,
S.H.Hughes,
E.Arnold.
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Ref.
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Nat Struct Biol, 1995,
2,
407-415.
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PubMed id
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Secondary reference #5
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Title
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Locations of anti-Aids drug binding sites and resistance mutations in the three-Dimensional structure of HIV-1 reverse transcriptase. Implications for mechanisms of drug inhibition and resistance.
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Authors
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C.Tantillo,
J.Ding,
A.Jacobo-Molina,
R.G.Nanni,
P.L.Boyer,
S.H.Hughes,
R.Pauwels,
K.Andries,
P.A.Janssen,
E.Arnold.
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Ref.
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J Mol Biol, 1994,
243,
369-387.
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PubMed id
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Secondary reference #6
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Title
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Buried surface analysis of HIV-1 reverse transcriptase p66/p51 heterodimer and its interaction with dsdna template/primer.
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Authors
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J.Ding,
A.Jacobo-Molina,
C.Tantillo,
X.Lu,
R.G.Nanni,
E.Arnold.
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Ref.
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J Mol Recognit, 1994,
7,
157-161.
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PubMed id
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Secondary reference #7
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Title
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Crystal structure of human immunodeficiency virus type 1 reverse transcriptase complexed with double-Stranded DNA at 3.0 a resolution shows bent DNA.
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Authors
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A.Jacobo-Molina,
J.Ding,
R.G.Nanni,
A.D.Clark,
X.Lu,
C.Tantillo,
R.L.Williams,
G.Kamer,
A.L.Ferris,
P.Clark.
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Ref.
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Proc Natl Acad Sci U S A, 1993,
90,
6320-6324.
[DOI no: ]
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PubMed id
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