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PDBsum entry 2nrt
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References listed in PDB file
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Key reference
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Title
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Structure of the c-Terminal half of uvrc reveals an rnase h endonuclease domain with an argonaute-Like catalytic triad.
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Authors
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E.Karakas,
J.J.Truglio,
D.Croteau,
B.Rhau,
L.Wang,
B.Van houten,
C.Kisker.
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Ref.
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EMBO J, 2007,
26,
613-622.
[DOI no: ]
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PubMed id
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Abstract
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Removal and repair of DNA damage by the nucleotide excision repair pathway
requires two sequential incision reactions, which are achieved by the
endonuclease UvrC in eubacteria. Here, we describe the crystal structure of the
C-terminal half of UvrC, which contains the catalytic domain responsible for 5'
incision and a helix-hairpin-helix-domain that is implicated in DNA binding.
Surprisingly, the 5' catalytic domain shares structural homology with RNase H
despite the lack of sequence homology and contains an uncommon DDH triad. The
structure also reveals two highly conserved patches on the surface of the
protein, which are not related to the active site. Mutations of residues in one
of these patches led to the inability of the enzyme to bind DNA and severely
compromised both incision reactions. Based on our results, we suggest a model of
how UvrC forms a productive protein-DNA complex to excise the damage from DNA.
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Figure 4.
Figure 4 Electrostatic surface potential and sequence
conservation of the C-terminal half of UvrC. (A) Electrostatic
surface potential was calculated with PyMol/APBS and contoured
at  10
k[B]T. The top panel features the active site of the protein and
the bottom view is a 180° rotation. (B) Sequence
conservation using the same orientations as in (A). The degree
of conservation was obtained by alignment of 47 UvrC sequences
with ClustalX. Strictly conserved (red), very highly conserved
(blue), highly conserved (green) and moderately conserved
(black) amino acids are highlighted. The remainder of the
protein is colored in gray. Bound sulfate molecules are shown in
all-bonds representation. Selected amino acids are labeled.
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Figure 6.
Figure 6 DNA binding model. (A) Side-by-side comparison of the
(HhH)[2] domain of RuvA (left), TmUvrC (center) and EcUvrC
(right) after superposition. The DNA backbone of the RuvA/DNA
complex (left panel) is shown as an orange worm. Selected
residues are shown in all-bonds representation and are labeled.
The N- and C-termini are indicated. The HhHI and HhHII motifs
are colored yellow and green, respectively. The helical linker
between the two motifs is colored blue. (B) The endonuclease
domains of eight TmUvrC^C-term structures are superimposed to
show the orientation of the (HhH)[2] domains relative to the
endonuclease domain in the different crystal forms. (C) Model of
TmUvrC interacting with DNA based on a superposition with the
Tn5 transposase–DNA complex. The endonuclease and (HhH)[2]
domain of TmUvrC are colored yellow and cyan, respectively. The
DNA is orange and drawn with spokes for clarity. The side chains
of the catalytic triad and D405 are depicted as all-bonds. The
bound magnesium is shown as a green sphere. In the left panel,
the (HhH)[2] domain is depicted in the position found in the
crystal structure. The DNA-interacting region of the (HhH)[2]
domain is shown in red. In the right panel, the (HhH)[2] domain
has been rotated to form a productive UvrC/DNA complex. A dashed
line indicates the connection point between the endonuclease
domain and the (HhH)[2] domain.
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The above figures are
reprinted
by permission from Macmillan Publishers Ltd:
EMBO J
(2007,
26,
613-622)
copyright 2007.
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