 |
PDBsum entry 1z96
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
 |
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
Protein transport
|
PDB id
|
|
|
|
1z96
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
References listed in PDB file
|
|
|
Key reference
|
|
|
Title
|
|
Mechanism of lys48-Linked polyubiquitin chain recognition by the mud1 uba domain.
|
|
|
Authors
|
|
J.F.Trempe,
N.R.Brown,
E.D.Lowe,
C.Gordon,
I.D.Campbell,
M.E.Noble,
J.A.Endicott.
|
|
|
Ref.
|
|
EMBO J, 2005,
24,
3178-3189.
[DOI no: ]
|
|
|
PubMed id
|
|
|
|
|
|
Abstract
|
|
|
The ubiquitin-pathway associated (UBA) domain is a 40-residue
polyubiquitin-binding motif. The Schizosaccharomyces pombe protein Mud1 is an
ortholog of the Saccharomyces cerevisiae DNA-damage response protein Ddi1 and
binds to K48-linked polyubiquitin through its UBA domain. We have solved the
crystal structure of Mud1 UBA at 1.8 angstroms resolution, revealing a canonical
three-helical UBA fold. We have probed the interactions of this domain using
mutagenesis, surface plasmon resonance, NMR and analytical ultracentrifugation.
We show that the ubiquitin-binding surface of Mud1 UBA extends beyond previously
recognized motifs and can be functionally dissected into primary and secondary
ubiquitin-binding sites. Mutation of Phe330 to alanine, a residue exposed
between helices 2 and 3, significantly reduces the affinity of the Mud1 UBA
domain for K48-linked polyubiquitin, despite leaving the primary binding surface
functionally intact. Moreover, K48-linked diubiquitin binds a single Mud1 UBA
domain even in the presence of excess UBA. We therefore propose a mechanism for
the recognition of K48-linked polyubiquitin chains by Mud1 in which diubiquitin
units are specifically recognized by a single UBA domain.
|
|
|
|
|
|
|
|
Figure 1.
Figure 1 Crystal structure of the UBA domain from Mud1. (A)
Cartoon representation of the asymmetric unit of the Mud1 UBA
crystal structure formed by chain A (red) and B (blue). Labeled
side chains are involved in domain -domain contacts or in Ub
binding, and are drawn in stick mode. The sulfur anomalous
difference map is shown as a blue contour at 10.0  .
(B) Electron density 2F[o] -F[c] map of Mud1 UBA crystal
structure at a contour level of 1.0 ,
showing a crystal contact between two 'A' monomers in the
crystal structure. Note the stacking of the side-chain phenyl
groups of two Phe330 residues. Residues in a different
symmetry-related molecule are tagged with an apostrophe.
|
|
Figure 7.
Figure 7 Molecular model for the interaction of Mud1 UBA with
K48-linked Ub[2]. (A) Intensity changes by cross-saturation of
the 15N -1H cross-peaks in 2H, 15N-Mud1 UBA in complex with
unlabeled K48-Ub[2]. (B) Primary (left) and secondary (right)
binding sites on Mud1UBA as identified by NMR cross-saturation.
Resonances showing intensity ratios <0.5 or 0.3 are displayed on
the molecular surface of Mud1 UBA in light or dark red,
respectively. (C) Closed conformation of Ub[2], based on the
crystal structure obtained under basic conditions (PDB accession
code 1AAR) (Cook et al, 1992). The hydrophobic patches on each
Ub moiety interact with each other. The proximal and distal
moieties of Ub[2] are colored in red and blue, respectively. (D)
Open conformation of Ub[2], in equilibrium with closed
conformation in solution. The two hydrophobic clusters formed by
residues Leu8, Ile44, His68 and Val70 are available for binding
of a single UBA domain via a primary (purple) and a secondary
(blue) Ub-binding sites.
|
|
|
|
|
|
The above figures are
reprinted
by permission from Macmillan Publishers Ltd:
EMBO J
(2005,
24,
3178-3189)
copyright 2005.
|
|
|
|
|
|
|
