PDBsum entry 1z96

Protein transport

PDB id: 1z96

Contents

Protein chains

38 a.a.

35 a.a.

Waters ×95

PDB id:

1z96

Name:

Protein transport

Title:

Crystal structure of the mud1 uba domain

Structure:

Uba-domain protein mud1. Chain: a, b. Fragment: c-terminal uba. Synonym: uba-domain containing protein 1. DNA-damage inducible protein ddi1 homologprotein mud1. Engineered: yes

Source:

Schizosaccharomyces pombe. Fission yeast. Organism_taxid: 4896. Gene: mud1. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008.

Biol. unit:

Monomer (from PQS)

Resolution:

1.80Å R-factor: 0.168 R-free: 0.196

Authors:

J.-F.Trempe,N.R.Brown,E.D.Lowe,M.E.M.Noble,C.Gordon,I.D.Campbell, L.N.Johnson,J.A.Endicott

Key ref:

J.F.Trempe et al. (2005). Mechanism of Lys48-linked polyubiquitin chain recognition by the Mud1 UBA domain. EMBO J, 24, 3178-3189. PubMed id: 16138082 DOI: 10.1038/sj.emboj.7600797

Date:

31-Mar-05 Release date: 04-Oct-05

Protein chain

Q10256  (MUD1_SCHPO) -  UBA domain-containing protein Mud1 from Schizosaccharomyces pombe (strain 972 / ATCC 24843)

Seq: 332 a.a.

Struc: 38 a.a.

Protein chain

Q10256  (MUD1_SCHPO) -  UBA domain-containing protein Mud1 from Schizosaccharomyces pombe (strain 972 / ATCC 24843)

Seq: 332 a.a.

Struc: 35 a.a.

Enzyme reactions

• Enzyme class: Chains A, B: E.C.3.4.23.- - ?????

DOI no: 10.1038/sj.emboj.7600797

EMBO J 24:3178-3189 (2005)

PubMed id: 16138082

Mechanism of Lys48-linked polyubiquitin chain recognition by the Mud1 UBA domain.

J.F.Trempe, N.R.Brown, E.D.Lowe, C.Gordon, I.D.Campbell, M.E.Noble, J.A.Endicott.

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.

Selected figure(s)

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.

Figures were selected by the author.