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PDBsum entry 1ua0
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Transferase/DNA
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PDB id
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1ua0
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Contents |
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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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Observing translesion synthesis of an aromatic amine DNA adduct by a high-Fidelity DNA polymerase.
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Authors
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G.W.Hsu,
J.R.Kiefer,
D.Burnouf,
O.J.Becherel,
R.P.Fuchs,
L.S.Beese.
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Ref.
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J Biol Chem, 2004,
279,
50280-50285.
[DOI no: ]
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PubMed id
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Abstract
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Aromatic amines have been studied for more than a half-century as model
carcinogens representing a class of chemicals that form bulky adducts to the C8
position of guanine in DNA. Among these guanine adducts, the
N-(2'-deoxyguanosin-8-yl)-aminofluorene (G-AF) and
N-2-(2'-deoxyguanosin-8-yl)-acetylaminofluorene (G-AAF) derivatives are the best
studied. Although G-AF and G-AAF differ by only an acetyl group, they exert
different effects on DNA replication by replicative and high-fidelity DNA
polymerases. Translesion synthesis of G-AF is achieved with high-fidelity
polymerases, whereas replication of G-AAF requires specialized bypass
polymerases. Here we have presented structures of G-AF as it undergoes one round
of accurate replication by a high-fidelity DNA polymerase. Nucleotide
incorporation opposite G-AF is achieved in solution and in the crystal,
revealing how the polymerase accommodates and replicates past G-AF, but not
G-AAF. Like an unmodified guanine, G-AF adopts a conformation that allows it to
form Watson-Crick hydrogen bonds with an opposing cytosine that results in
protrusion of the bulky fluorene moiety into the major groove. Although
incorporation opposite G-AF is observed, the C:G-AF base pair induces
distortions to the polymerase active site that slow translesion synthesis.
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Figure 1.
FIG. 1. A, chemical structures of G-AF and G-AAF. dR,
deoxyribose ring. The proton and the N-acetyl group that
differentiate G-AF from G-AAF are shown in orange. B, schematic
of the polymerase active site. Sites through which the template
base (red) traverses during replication are shown (see the
Introduction for description).
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Figure 4.
FIG. 4. Structure of BF with G-AF at the postinsertion
site. A, structure of BF bound to G-AF at the postinsertion site
(blue) superimposed with a structure of BF bound to an
unmodified base at the postinsertion site (gray). G-AF (red)
adopts an anti conformation and obstructs the n+1 template base
from occupying the preinsertion site that is itself disordered.
B, C:G-AF base pair surrounded by electron density contoured at
3.5  and calculated using
Fourier coefficients (F[obs] - F[calc])  [calc] with C:G-AF
omitted from the final model. Hydrogen bonds (dashed lines) are
shown accompanied with bond lengths.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2004,
279,
50280-50285)
copyright 2004.
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Secondary reference #1
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Title
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Structures of mismatch replication errors observed in a DNA polymerase.
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Authors
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S.J.Johnson,
L.S.Beese.
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Ref.
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Cell, 2004,
116,
803-816.
[DOI no: ]
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PubMed id
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Figure 2.
Figure 2. DNA Mismatches Bound at the Polymerase
Postinsertion SiteThe bases are shown in the same orientation
and location as the G•C base pair in Figure 1B. Left, hydrogen
bonding pattern. Right, superimposition of the molecular surface
of the mismatch (red) and cognate G•C base pair (yellow, PDB
ID 1L3S) bound at the postinsertion site, highlighting
differences in shape and location of the primer terminus.
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Figure 4.
Figure 4. Extension of a G•T Mismatch by Successive
Rounds of ReplicationThe conformation of the G•T mismatch is
shown at each position (left), including interacting water
molecules (red spheres). Dashed lines indicate potential
hydrogen bonds. At the n-3 and n-4 positions, hydrogen bonds are
shown between groups within the appropriate distance (≤3.2
Å) and correspond to tautomerization or ionization of one
of the bases (see text). A schematic representation (right) of
the mismatch complex, drawn and color coded as described in
Figure 1, Figure 2 and Figure 3, indicates regions of the
polymerase active site that are disrupted upon binding of the
mismatch (red line). Mismatch binding at positions n-1 to n-4
along the DNA duplex binding region (gray) results in a
distorted open conformation at the polymerase active site as
described by mechanism 1 (Figure 3). The normal open
conformation observed with homoduplexes is fully restored when
the mismatch is bound at the n-6 position.
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The above figures are
reproduced from the cited reference
with permission from Cell Press
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Secondary reference #2
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Title
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Processive DNA synthesis observed in a polymerase crystal suggests a mechanism for the prevention of frameshift mutations.
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Authors
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S.J.Johnson,
J.S.Taylor,
L.S.Beese.
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Ref.
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Proc Natl Acad Sci U S A, 2003,
100,
3895-3900.
[DOI no: ]
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PubMed id
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Figure 2.
Fig. 2. Active site superposition of open and closed BF
structures. (A) The 11-bp open binary complex (yellow) and the
closed ternary complex (blue) are shown in stereo view. The
largest conformational differences occur in the fingers domain,
including the O helix, O1 helix, and preinsertion site. The
acceptor template base (n) occupies the preinsertion site in the
open conformation and the insertion site in the closed
conformation. (B) A close-up view of the preinsertion site. The
locations of the conserved Tyr-714 are indicated.
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Figure 3.
Fig. 3. Conformational interlocks during DNA synthesis. A
schematic overview of the polymerase active site (A) and atomic
coordinates (B) derived from the open and closed BF structures
represent a complete round of DNA synthesis. The conformational
changes described here are presented in animated form in Movie
1, which is published as supporting information on the PNAS web
site. The reaction cycle starts with the acceptor template base
(n, red) bound at the template preinsertion site (between the O
and O1 helices; green shading); Tyr-714 blocks access to the
insertion site (blue shading) and stacks with the n-1 base pair
at the postinsertion site (gray shading). Formation of the
closed conformation involves rearrangement of the O and O1
helices, which simultaneously blocks the template preinsertion
site and unblocks the insertion site. These rearrangements move
the acceptor template base (n) to the insertion site, where it
pairs with an incoming dNTP (green). Nucleotide incorporation
occurs on formation of a cognate base pair and proper assembly
of the catalytic site (orange shading). The cycle is completed
with translocation of the DNA by one base pair position. The
polymerase resets to the open conformation in preparation for
the next round of DNA synthesis.
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Secondary reference #3
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Title
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Visualizing DNA replication in a catalytically active bacillus DNA polymerase crystal.
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Authors
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J.R.Kiefer,
C.Mao,
J.C.Braman,
L.S.Beese.
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Ref.
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Nature, 1998,
391,
304-307.
[DOI no: ]
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PubMed id
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Figure 1.
Figure 1 Structure of the Bacillus fragment with duplex DNA
bound at the polymerase active site. The Bacillus fragment
molecular surface is coloured according to its proximity to the
DNA, with all points less than 3.5 ? coloured magenta, between
3.5 and 5.0 ? yellow, and greater than 5 ? blue. Bound water
molecules were not included in this calculation.
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Figure 4.
Figure 4 Polymerase active site with observed DNA and
modelled dTTP. The position of dTTP (violet) was based on the
 -polymerase
complex18, adjusted such that the base ring stacks with the
primer and one oxygen from each phosphate group was within 3 ?
of the observed metal ion (gold). The sugar pucker of the primer
terminus was made C3'-endo, which shifted its 3'-OH to within
1.7 ? of the modelled  -phosphate
of the dTTP. A second metal ion (violet) was modelled to be
within 3 ? of the 3'-OH of the primer, the -phosphate
group, and residues Asp 830 and Glu 831. The observed 5'
template overhang cannot accept an incoming dNTP without a
conformational change of the O helix.
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The above figures are
reproduced from the cited reference
with permission from Macmillan Publishers Ltd
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