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PDBsum entry 1nwq
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Transcription/DNA
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PDB id
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1nwq
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Contents |
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* Residue conservation analysis
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References listed in PDB file
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Key reference
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Title
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Structural basis for DNA recognition by the basic region leucine zipper transcription factor ccaat/enhancer-Binding protein alpha.
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Authors
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M.Miller,
J.D.Shuman,
T.Sebastian,
Z.Dauter,
P.F.Johnson.
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Ref.
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J Biol Chem, 2003,
278,
15178-15184.
[DOI no: ]
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PubMed id
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Abstract
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CCAAT/enhancer-binding proteins (C/EBPs) are basic region leucine zipper (bZIP)
transcription factors that regulate cell differentiation, growth, survival, and
inflammation. To understand the molecular basis of DNA recognition by the C/EBP
family we determined the x-ray structure of a C/EBPalpha bZIP polypeptide bound
to its cognate DNA site (A(-5)T(-4)T(-3)G(-2)C(-1)G(1)C(2)A(3)A(4)T(5)) and
characterized several basic region mutants. Binding specificity is provided by
interactions of basic region residues Arg(289), Asn(292), Ala(295), Val(296),
Ser(299), and Arg(300) with DNA bases. A striking feature of the C/EBPalpha
protein-DNA interface that distinguishes it from known bZIP-DNA complexes is the
central role of Arg(289), which is hydrogen-bonded to base A(3), phosphate,
Asn(292) (invariant in bZIPs), and Asn(293). The conformation of Arg(289) is
also restricted by Tyr(285). In accordance with the structural model, mutation
of Arg(289) or a pair of its interacting partners (Tyr(285) and Asn(293))
abolished C/EBPalpha binding activity. Val(296) (Ala in most other bZIPs)
contributes to C/EBPalpha specificity by discriminating against purines at
position -3 and imposing steric restraints on the invariant Arg(300). Mutating
Val(296) to Ala strongly enhanced C/EBPalpha binding to cAMP response element
(CRE) sites while retaining affinity for C/EBP sites. Thus, Arg(289) is
essential for formation of the complementary protein-DNA interface, whereas
Val(296) functions primarily to restrict interactions with related sequences
such as CRE sites rather than specifying binding to C/EBP sites. Our studies
also help to explain the phenotypes of mice carrying targeted mutations in the
C/EBPalpha bZIP region.
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Figure 4.
Fig. 4. Architecture of the protein surface complementary
to the cognate DNA. A, critical interactions in the C/EBP  protein-DNA
interface. Protein side chains are represented as sticks and DNA
as balls-and-sticks. Selected electrostatic and van der Waals
interactions are depicted as dashed and dotted lines,
respectively. B, comparison of conformations of the conserved
side chains in the basic regions of C/EBP (green) and
GCN4 (gray). The side chain of the invariant Asn residue from
the PAP1 structure is shown in yellow.
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Figure 5.
Fig. 5. Details of the C/EBP protein-DNA
interface. A, hydrophobic cluster involving two thymine moieties
from the C/EBP site and side chains of Val296 and Arg300. B,
interactions defining possible conformation of Arg300. The side
chain of the homologous Arg residue from GCN4 (one observed
conformation) is shown in gray. C, comparison of the protein
environment of base 2 in C/EBP (green) and
GCN4 (gray) complexes. Hydrophobic interaction of the methyl
group from T2 and Ala occurring in GCN4 is marked by a dotted
line. Note that C^2 in the C/EBP site does not interact with
Val296, and its position is displaced relative to T2 from the
GCN4-CREB complex. D, electron density (2F[o]  F[c])
contoured at 1.0  (gray) and
1.8 (green;
DNA only) is shown for important residues, with the coordinates
superimposed.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2003,
278,
15178-15184)
copyright 2003.
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