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PDBsum entry 1svx
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De novo protein/sugar binding protein
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PDB id
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1svx
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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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High-Affinity binders selected from designed ankyrin repeat protein libraries.
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Authors
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H.K.Binz,
P.Amstutz,
A.Kohl,
M.T.Stumpp,
C.Briand,
P.Forrer,
M.G.Grütter,
A.Plückthun.
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Ref.
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Nat Biotechnol, 2004,
22,
575-582.
[DOI no: ]
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PubMed id
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Abstract
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We report here the evolution of ankyrin repeat (AR) proteins in vitro for
specific, high-affinity target binding. Using a consensus design strategy, we
generated combinatorial libraries of AR proteins of varying repeat numbers with
diversified binding surfaces. Libraries of two and three repeats, flanked by
'capping repeats,' were used in ribosome-display selections against maltose
binding protein (MBP) and two eukaryotic kinases. We rapidly enriched
target-specific binders with affinities in the low nanomolar range and
determined the crystal structure of one of the selected AR proteins in complex
with MBP at 2.3 A resolution. The interaction relies on the randomized positions
of the designed AR protein and is comparable to natural, heterodimeric
protein-protein interactions. Thus, our AR protein libraries are valuable
sources for binding molecules and, because of the very favorable biophysical
properties of the designed AR proteins, an attractive alternative to antibody
libraries.
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Figure 4.
Figure 4. Crystal structure of the designed AR protein off7 in
complex with MBP. (a,b) Two perpendicular views of the
complex are shown. MBP is on the left (blue), off7 on the right
(ochre). The interaction residues are highlighted in stick-mode
in red (off7) and blue (MBP), respectively. (c) A close stereo
view on the H-bond pattern in similar view as in b. Note that in
this representation only residues involved in H-bonds (green
dashed lines) are shown. For orientation, some residues involved
in H-bonding are labeled. Figure 4a -c were made with MolMol49.
(d) Ligplot48 representation of the interaction between MBP
(chain B, blue) and off7 (chain A, red). H-bonds (in green)
including the H-bond distances as well as residues and atoms
involved in hydrophobic contacts (indicated by red or blue rays)
are shown.
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Figure 5.
Figure 5. Open sandwich illustrations of the interaction
surfaces of AR proteins and their targets. (a -c) GRASP50
shape complementarity representations of the interactions
between off7 and MBP (shape complementarity, 0.739) (a), GABP
 1
and GABP  (PDB
entry 1AWC; shape complementarity, 0.665) (b), and p18^INK4c and
CDK6 (PDB entry 1G3N; shape complementarity, 0.688) (c),
respectively. The complex is shown on the left with the AR
proteins in a backbone worm representation ( -helices
in blue, -turns
in green) and the target protein in a surface representation.
The open sandwich surface representations are shown in the
middle (AR proteins) and on the right (targets). The contact
areas are stained according to the shape complementarities from
orange (low) to red (high).
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The above figures are
reprinted
by permission from Macmillan Publishers Ltd:
Nat Biotechnol
(2004,
22,
575-582)
copyright 2004.
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