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PDBsum entry 1fxt
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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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Structure of a conjugating enzyme-Ubiquitin thiolester intermediate reveals a novel role for the ubiquitin tail.
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Authors
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K.S.Hamilton,
M.J.Ellison,
K.R.Barber,
R.S.Williams,
J.T.Huzil,
S.Mckenna,
C.Ptak,
M.Glover,
G.S.Shaw.
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Ref.
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Structure, 2001,
9,
897-904.
[DOI no: ]
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PubMed id
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Abstract
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BACKGROUND: Ubiquitin-conjugating enzymes (E2s) are central enzymes involved in
ubiquitin-mediated protein degradation. During this process, ubiquitin (Ub) and
the E2 protein form an unstable E2-Ub thiolester intermediate prior to the
transfer of ubiquitin to an E3-ligase protein and the labeling of a substrate
for degradation. A series of complex interactions occur among the target
substrate, ubiquitin, E2, and E3 in order to efficiently facilitate the transfer
of the ubiquitin molecule. However, due to the inherent instability of the E2-Ub
thiolester, the structural details of this complex intermediate are not known.
RESULTS: A three-dimensional model of the E2-Ub thiolester intermediate has been
determined for the catalytic domain of the E2 protein Ubc1 (Ubc1(Delta450)) and
ubiquitin from S. cerevisiae. The interface of the E2-Ub intermediate was
determined by kinetically monitoring thiolester formation by 1H-(15)N HSQC
spectra by using combinations of 15N-labeled and unlabeled Ubc1(Delta450) and Ub
proteins. By using the surface interface as a guide and the X-ray structures of
Ub and the 1.9 A structure of Ubc1(Delta450) determined here, docking
simulations followed by energy minimization were used to produce the first model
of a E2-Ub thiolester intermediate. CONCLUSIONS: Complementary surfaces were
found on the E2 and Ub proteins whereby the C terminus of Ub wraps around the E2
protein terminating in the thiolester between C88 (Ubc1(Delta450)) and G76 (Ub).
The model supports in vivo and in vitro experiments of E2 derivatives carrying
surface residue substitutions. Furthermore, the model provides insights into the
arrangement of Ub, E2, and E3 within a ternary targeting complex.
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Figure 3.
Figure 3. Model of the E2-Ub Thiolester IntermediateThe
model was determined by Monte Carlo docking as described in
Experimental Procedures. The model shows (a) side and (b) end
orientation views of helix a2 in the E2 molecule. Residues are
indicated on both E2 and Ub to indicate important side
chain-side chain interactions that arise at the protein-protein
interface, as described in the text. In both figures, the
thiol-forming C88 residue in E2 is a shown as a yellow
ball-and-stick representation near the (a) center and (b) top of
the complex
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The above figure is
reprinted
by permission from Cell Press:
Structure
(2001,
9,
897-904)
copyright 2001.
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