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PDBsum entry 2ca6
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Signaling regulator
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PDB id
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2ca6
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Contents |
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* Residue conservation analysis
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DOI no:
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Acta Crystallogr D Biol Crystallogr
62:750-765
(2006)
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PubMed id:
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Detecting and overcoming hemihedral twinning during the MIR structure determination of Rna1p.
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R.C.Hillig,
L.Renault.
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ABSTRACT
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The structure of Rna1p was originally solved to 2.7 A resolution by MIRAS from
crystals with partial hemihedral twinning in space group I4(1) [Hillig et al.
(1999), Mol. Cell, 3, 781-791] by finding a low-twinned native crystal (twin
fraction alpha=0.06) and after twin correction of all data sets. Rna1p crystals
have now been used to examine how far twinning and twin correction affect MIR
phasing with a higher resolution but highly twinned native data set. Even high
hemihedral twinning [alphanative=0.39, alphaderivative=0.24] would not have
hindered heavy-atom site identification of strong derivatives using difference
Patterson maps. However, a weaker derivative could have been missed and
refinement would have stalled at high R values had twinning not been identified
and accounted for. Twin correction improved both site identification,
experimental phasing statistics and MIR map quality. Different strategies were
tested for refinement against twinned data. Using uncorrected twinned data and
TWIN-CNS, Rna1p has now been refined to 2.2 A resolution (final twinned R and
Rfree were 0.165 and 0.218, respectively). The increased resolution enabled
release of the NCS restraints and allowed new conclusions to be drawn on the
flexibility of the two molecules in the asymmetric unit. In the case of Rna1p,
twinned crystal growth was possible owing to the presence of a twofold NCS axis
almost parallel to the twin operator.
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Selected figure(s)
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Figure 1.
Figure 1 Crystals and diffraction pattern of S. pombe Rna1p.
(a) Typical sea-urchin-like crystal clusters. (b) Manually
separated crystals of about 600 × 40 × 40 µm.
(c) Diffraction pattern from native-A (high-resolution sweep,
  =
0.5°, exposure time 8 min, detector edge 2 Å). Yellow
boxes denote the enlarged regions. The reflections show no signs
of splitting.
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Figure 6.
Figure 6 Structure of Rna1p refined to 2.20 Å. (a)
Representative view of the final 3F[o] - 2F[c] electron-density
map. Shown is the region around leucine-rich repeat 8 (LRR8),
contoured at 1.5  .
(b) Ribbon representation of Rna1p. (c) Superimposition of the
two independent molecules A (red) and B (blue) in the asymmetric
unit (C^  backbone
representation). Differences are found in the N-terminal region
as well as in LRR3/LRR4. (d) Superimposition of Rna1p (molecules
A and B in red and blue, respectively) and the complex of Rna1p
(green) with Ran-GMPPNP-Mg-RanBP1 (PDB code 1k5d ). The Ran
backbone is shown as a grey ribbon, GMPPNP in stick
representation and Mg as magenta-coloured sphere. An enlargement
of the region of the flexible loops of LRR3/LRR4 is shown. This
region, which differs between molecules A and B in the
high-resolution structure of Rna1p, coincides with part of the
interface between Rna1p and Ran-GMPPNP. The flexibility in Rna1p
may indicate an inherent mobility designed to allow an induced
fit.
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The above figures are
reprinted
by permission from the IUCr:
Acta Crystallogr D Biol Crystallogr
(2006,
62,
750-765)
copyright 2006.
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Figures were
selected
by an automated process.
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Links
