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PDBsum entry 1wvn
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RNA binding protein
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PDB id
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1wvn
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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 and RNA binding of the third kh domain of poly(c)-Binding protein 1.
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Authors
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M.Sidiqi,
J.A.Wilce,
J.P.Vivian,
C.J.Porter,
A.Barker,
P.J.Leedman,
M.C.Wilce.
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Ref.
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Nucleic Acids Res, 2005,
33,
1213-1221.
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PubMed id
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Abstract
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Poly(C)-binding proteins (CPs) are important regulators of mRNA stability and
translational regulation. They recognize C-rich RNA through their triple KH (hn
RNP K homology) domain structures and are thought to carry out their function
though direct protection of mRNA sites as well as through interactions with
other RNA-binding proteins. We report the crystallographically derived structure
of the third domain of alphaCP1 to 2.1 A resolution. alphaCP1-KH3 assumes a
classical type I KH domain fold with a triple-stranded beta-sheet held against a
three-helix cluster in a betaalphaalphabetabetaalpha configuration. Its binding
affinity to an RNA sequence from the 3'-untranslated region (3'-UTR) of androgen
receptor mRNA was determined using surface plasmon resonance, giving a K(d) of
4.37 microM, which is indicative of intermediate binding. A model of
alphaCP1-KH3 with poly(C)-RNA was generated by homology to a recently reported
RNA-bound KH domain structure and suggests the molecular basis for
oligonucleotide binding and poly(C)-RNA specificity.
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Figure 1.
The crystal structure of
Å resolution depicted in (A) cartoon form and (B) as a molecular surface in the same
orientation. The structure is shown from the beginning of ß-strand 1 to the end of
{alpha} -helix 3, since the regions outside these bounds were random coil or not
visible in the density. The GXXG motif, common to this oligonucleotide-binding motif, is
coloured blue. The `variable loop' region between ß-sheets 2 and 3 is coloured pink. These
regions bound the hydrophobic oligonucleotide-binding cleft that accommodates C-rich RNA
or ssDNA. (C) The electrostatic potential emanating from the {alpha} CP1-KH3
structure calculated using the APBS software package (http://agave.wustl.edu/apbs/)
(39-43). Potential contours are shown at +1 kT/e (blue) and -1 kT/e (red) and
obtained by solution of the linearized Poisson-Boltzmann equation at 150 mM ionic strength
with a solute dielectric of 2 and a solvent dielectric of 78.5. The blue contour
represents a striking positive potential directing oligonucleotides to the binding cleft.
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Figure 4.
Interaction of the
measured by surface plasmon resonance. (A) 30 RU RNA was immobilized on a
streptavidin-coated chip. Binding interactions were measured for a series of dilutions of
the {alpha} CP1-KH3 domain from 10 to 0.625 µM for 2 min using flow rate of 50
µl/min. (B) Steady-state analysis of the interaction yielded a K[d] value of 4.37 µM.
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The above figures are
reprinted
from an Open Access publication published by Oxford University Press:
Nucleic Acids Res
(2005,
33,
1213-1221)
copyright 2005.
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