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PDBsum entry 3how
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Transcription,transferase/DNA/RNA hybrid
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PDB id
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3how
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Contents |
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1416 a.a.
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1112 a.a.
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266 a.a.
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178 a.a.
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214 a.a.
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87 a.a.
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171 a.a.
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134 a.a.
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114 a.a.
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65 a.a.
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114 a.a.
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44 a.a.
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References listed in PDB file
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Key reference
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Title
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Structural basis of transcription: mismatch-Specific fidelity mechanisms and paused RNA polymerase ii with frayed RNA.
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Authors
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J.F.Sydow,
F.Brueckner,
A.C.Cheung,
G.E.Damsma,
S.Dengl,
E.Lehmann,
D.Vassylyev,
P.Cramer.
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Ref.
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Mol Cell, 2009,
34,
710-721.
[DOI no: ]
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PubMed id
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Abstract
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We show that RNA polymerase (Pol) II prevents erroneous transcription in vitro
with different strategies that depend on the type of DNARNA base mismatch.
Certain mismatches are efficiently formed but impair RNA extension. Other
mismatches allow for RNA extension but are inefficiently formed and efficiently
proofread by RNA cleavage. X-ray analysis reveals that a TU mismatch impairs RNA
extension by forming a wobble base pair at the Pol II active center that
dissociates the catalytic metal ion and misaligns the RNA 3' end. The mismatch
can also stabilize a paused state of Pol II with a frayed RNA 3' nucleotide. The
frayed nucleotide binds in the Pol II pore either parallel or perpendicular to
the DNA-RNA hybrid axis (fraying sites I and II, respectively) and overlaps the
nucleoside triphosphate (NTP) site, explaining how it halts transcription during
proofreading, before backtracking and RNA cleavage.
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Figure 5.
Figure 5. Frayed Nucleotides Overlap the NTP, Closed Trigger
Loop, and the TFIIS Hairpin
(A) Frayed nucleotides overlap
the NTP bound to the insertion site (green cyan, taken from
bacterial Pol EC, PDB-code 2O5J [Vassylyev et al., 2007b]). Van
der Waals radii are represented by colored dots. All structures
were superimposed with their active site regions.
(B and C)
Frayed nucleotides overlap the closed trigger loop (cyan) at
residue F1084 (B, taken from the Pol II EC, PDB-code 2E2H [Wang
et al., 2006]) and/or at residue H1242 (C, bacterial Pol EC,
PDB-code 2O5J [Vassylyev et al., 2007b]).
(D) Frayed
nucleotides overlap the tip of the hairpin of the
cleavage-stimulatory factor TFIIS. The structures of EC III, EC
IV, and the Pol II-TFIIS complex (PDB-code 1PQV, [Kettenberger
et al., 2003]) were superimposed with their active center
regions. TFIIS is shown in orange. The canonical side view is
used.
(E) Detailed view of the superposition in (D) around
the active site, revealing a potential clash of the TFIIS acidic
hairpin with the frayed nucleotides.
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Figure 6.
Figure 6. Fork Loop 2-Downstream DNA Contact
(A)
Comparison of the conformation of fork loop 2 in EC III with
that in previous Pol II EC structures (PDB-codes 1Y1W
(Kettenberger et al., 2004) and 2E2I (Wang et al., 2006).
(B) Interaction of the side chain of fork loop 2 Rpb2 residue
R504 with the guanine base at position +4 of downstream DNA. The
final 2F[o]-F[c] electron density is shown in blue, contoured at
0.7 σ.
(C) Interaction of regions in EC III that may be
involved in pausing, including the frayed nucleotide, βDloopII,
the bridge helix, fork loop 2, and downstream DNA.
(D)
Multiple sequence alignment of fork loop 2 and surrounding Rpb2
residues from S. cerevisiae, H. sapiens, P. furiosus, E. coli,
and T. thermophilus (CLUSTAL W). The conserved R504 from S.
cerevisiae is highlighted in blue.
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The above figures are
reprinted
by permission from Cell Press:
Mol Cell
(2009,
34,
710-721)
copyright 2009.
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