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PDBsum entry 1qzh
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DNA binding protein/DNA
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PDB id
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1qzh
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References listed in PDB file
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Key reference
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Title
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Dna self-Recognition in the structure of pot1 bound to telomeric single-Stranded DNA.
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Authors
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M.Lei,
E.R.Podell,
P.Baumann,
T.R.Cech.
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Ref.
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Nature, 2003,
426,
198-203.
[DOI no: ]
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PubMed id
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Abstract
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Telomeres, specialized protein-DNA complexes that cap the ends of linear
chromosomes, are essential for protecting chromosomes from degradation and
end-to-end fusions. The Pot1 (protection of telomeres 1) protein is a widely
distributed eukaryotic end-capping protein, having been identified in fission
yeast, microsporidia, plants and animals. Schizosaccharomyces pombe Pot1p is
essential for telomere maintenance, and human POT1 has been implicated in
telomerase regulation. Pot1 binds telomeric single-stranded DNA (ssDNA) with
exceptionally high sequence specificity, the molecular basis of which has been
unknown. Here we describe the 1.9-A-resolution crystal structure of the
amino-terminal DNA-binding domain of S. pombe Pot1p complexed with ssDNA. The
protein adopts an oligonucleotide/oligosaccharide-binding (OB) fold with two
loops that protrude to form a clamp for ssDNA binding. The structure explains
the sequence specificity of binding: in the context of the Pot1 protein, DNA
self-recognition involving base-stacking and unusual G-T base pairs compacts the
DNA. Any sequence change disrupts the ability of the DNA to form this structure,
preventing it from contacting the array of protein hydrogen-bonding groups. The
structure also explains how Pot1p avoids binding the vast excess of RNA in the
nucleus.
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Figure 2.
Figure 2: Protein -ssDNA and ssDNA -ssDNA interactions in the
Pot1pN -GGTTAC complex. Potp1N is shown in blue, and the
ssDNA is coloured as in Fig. 1a. Protein -DNA intermolecular
hydrogen bonds are shown as dotted green lines, and ssDNA
intramolecular hydrogen bonds as dotted yellow lines. All panels
except e were generated using Molscript and Raster 3D (refs 28,
29). a, Stereoimage of the protein -ssDNA interactions. The
ssDNA is bound in a compact and folded conformation. b, c, ssDNA
self-recognition by G -T base-pairing interactions. The base
pairs are oriented such that all Watson -Crick donor/acceptor
groups of the bases face the inner side of the binding groove
and make extensive hydrogen-bonding interactions with the
protein. d, Interactions of the 3' end of the ssDNA. The
hydrogen bonds between A5 and the backbone phosphodiester groups
of T3 and T4 represent the second form of self-recognition. e,
Schematic representation of the Pot1pN -ssDNA interactions.
Bases of the DNA are shown as purple bars; phosphodiester groups
as yellow circles; sugar rings as cyan pentamers; and protein
residues as green boxes. Stacking and van der Waals interactions
between bases and protein are shown as blue arrows, and stacking
between bases as red arrows. Base-paring interactions of the
ssDNA are shown as red dashed lines.
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Figure 3.
Figure 3: Mutational analysis of residues important for Pot1pN
-ssDNA interaction. The protein is shown in blue and the DNA
in yellow. The mutated residues are shown in a ball-and-stick
model. Red, non-functional mutations T62V (0/84) and F88A
(0/156); green, functional mutations E13A (15/24), G47A
(88/160), I48T (62/148), T111V (24/48) and K153A (17/36). The
numbers in parentheses indicate the results of the
complementation analysis (number of isolates that lack wild-type
pot1^+ versus total number of isolates tested).
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The above figures are
reprinted
by permission from Macmillan Publishers Ltd:
Nature
(2003,
426,
198-203)
copyright 2003.
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