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PDBsum entry 2qho
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Protein binding/ligase
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PDB id
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2qho
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Contents |
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72 a.a.
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47 a.a.
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76 a.a.
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49 a.a.
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49 a.a.
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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 of ubiquitin recognition by the ubiquitin-Associated (uba) domain of the ubiquitin ligase edd.
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Authors
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G.Kozlov,
L.Nguyen,
T.Lin,
G.De crescenzo,
M.Park,
K.Gehring.
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Ref.
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J Biol Chem, 2007,
282,
35787-35795.
[DOI no: ]
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PubMed id
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Abstract
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EDD (or HYD) is an E3 ubiquitin ligase in the family of HECT (homologous to
E6-AP C terminus) ligases. EDD contains an N-terminal ubiquitin-associated (UBA)
domain, which is present in a variety of proteins involved in ubiquitin-mediated
processes. Here, we use isothermal titration calorimetry (ITC), NMR titrations,
and pull-down assays to show that the EDD UBA domain binds ubiquitin. The 1.85 A
crystal structure of the complex with ubiquitin reveals the structural basis of
ubiquitin recognition by UBA helices alpha1 and alpha3. The structure shows a
larger number of intermolecular hydrogen bonds than observed in previous
UBA/ubiquitin complexes. Two of these involve ordered water molecules. The
functional importance of residues at the UBA/ubiquitin interface was confirmed
using site-directed mutagenesis. Surface plasmon resonance (SPR) measurements
show that the EDD UBA domain does not have a strong preference for polyubiquitin
chains over monoubiquitin. This suggests that EDD binds to monoubiquitinated
proteins, which is consistent with its involvement in DNA damage repair pathways.
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Figure 1.
FIGURE 1. The UBA domain of EDD binds to ubiquitin and
polyubiquitin. A, in vitro binding of the EDD and Cbl-b UBA
domains to ubiquitin agarose. B, in vitro binding of the EDD and
Cbl-b UBA domain to ubiquitin chains. C, calorimetric titration
of the EDD UBA domain with ubiquitin. The panel shows the
integrated heat released after correction for the heat of
dilution (data points, squares) and the curve of best fit for
binding to a single site.
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Figure 4.
FIGURE 4. Crystal structure of the EDD UBA-ubiquitin
complex. A, overall structure of the complex. Ubiquitin is in
yellow, and the UBA domain is in purple. B, UBA Val^196 binds
the hydrophobic groove formed by Ile^44, His^68, and Val^70 of
ubiquitin. C, UBA Leu^224 binds the groove formed by Ile^44,
Arg^42, Gln^49, and Val^70 of ubiquitin. D, Arg^42 and Gln^49 of
ubiquitin form a network of intermolecular hydrogen bonds with
Asn^221 and Ser^225 of EDD UBA. E, amide of Gly^47 of ubiquitin
forms an intermolecular hydrogen bond with carbonyl of Val^216
of EDD UBA via a bound water molecule. F, bound water molecule
facilitates hydrogen bonding between amide of Leu^71 of
ubiquitin and side chain of Asn^217 of EDD UBA. The 2F[o]-F[c]
electron density (1  contour, omit map) for
bound water molecules is shown in panels E and F.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2007,
282,
35787-35795)
copyright 2007.
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