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PDBsum entry 2v1c
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Recombination
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PDB id
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2v1c
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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 structure and mutational study of recor provide insight into its mode of DNA binding.
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Authors
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J.Timmins,
I.Leiros,
S.Mcsweeney.
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Ref.
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EMBO J, 2007,
26,
3260-3271.
[DOI no: ]
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PubMed id
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Abstract
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The crystal structure of the complex formed between Deinococcus radiodurans RecR
and RecO (drRecOR) has been determined. In accordance with previous biochemical
characterisation, the drRecOR complex displays a RecR:RecO molecular ratio of
2:1. The biologically relevant drRecOR entity consists of a heterohexamer in the
form of two drRecO molecules positioned on either side of the tetrameric ring of
drRecR, with their OB (oligonucleotide/oligosaccharide-binding) domains pointing
towards the interior of the ring. Mutagenesis studies validated the
protein-protein interactions observed in the crystal structure and allowed
mapping of the residues in the drRecOR complex required for DNA binding.
Furthermore, the preferred DNA substrate of drRecOR was identified as being
3'-overhanging DNA, as encountered at ssDNA-dsDNA junctions. Together these
results suggest a possible mechanism for drRecOR recognition of stalled
replication forks.
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Figure 3.
Figure 3 Study of the ionic interactions between drRecO and
drRecR. (A) Ribbon illustration of a monomer and a tetramer of
drRecR with ball-and-sticks representation of the mutated
residues. (B) Ribbon illustration of drRecO with ball-and-sticks
representation of the mutated residues. Residues coloured in
blue were mutated in order to disrupt protein–protein
interactions, whereas residues in red were predicted to be
involved in protein–DNA contacts. (C) Overlay of the
chromatograms obtained when purifying wild-type drRecOR (C1) and
mutant drRecOR (C2) on a Superdex 200 size-exclusion column. The
three peaks are labelled from 1 to 3. (D) Western blot analysis
of fractions corresponding to peaks 1, 2 and 3 from the gel
filtration runs of each of the drRecOR complexes (C1–C7 and
C10–C16). The Western blots were duplicated and were stained
for either drRecO (upper bands) or drRecR (lower bands). (E)
Illustration of the drRecR (gold)—drRecO (blue) interface with
a ball-and-sticks representation of drRecO-His93 and
drRecR-Glu146, displaying the 2mFo-DFc sigmaA-weighted electron
density map contoured at 1.3  .
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Figure 6.
Figure 6 Model for RecOR recognition of stalled replication
forks. Whereas the role of RecF in this process is still
unclear, it is known to associate with DNA in an ATP-dependent
fashion. Upon binding of RecOR to ssDNA–dsDNA junctions (step
2), we propose that interactions with RecF, SSB and/or DNA may
cause a structural rearrangement of RecOR (e.g. one RecO and two
RecR molecules may dissociate from the RecOR complex).
RecF-dependent ATP hydrolysis may provide the necessary energy
for this reorganisation (step 3) resulting in the formation of a
stable complex between RecOR and the stalled replication fork
(step 4). As a consequence, the assembled RecOR complex may
initiate the displacement of SSB and thus facilitate the loading
of RecA onto ssDNA, allowing for homologous recombination to
take place (step 5).
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The above figures are
reprinted
by permission from Macmillan Publishers Ltd:
EMBO J
(2007,
26,
3260-3271)
copyright 2007.
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