PDBsum entry 2khh

Transport protein

PDB id: 2khh

Contents

Protein chain

57 a.a. *

Ligands

ASP-SER-GLY-PHE-SER-PHE-GLY-SER-LYS

* Residue conservation analysis

PDB id:

2khh

Name:

Transport protein

Title:

Structural requirements for the uba domain of the mRNA export factor mex67 to bind its specific targets, the transcription elongation tho complex component hpr1 and nucleoporin fxfg repeats

Structure:

mRNA export factor mex67. Chain: a. Fragment: tap-c domain, unp residues 542-599. Engineered: yes. Fxfg. Chain: b. Engineered: yes

Source:

Saccharomyces cerevisiae. Yeast. Organism_taxid: 4932. Gene: mex67. Expressed in: escherichia coli. Expression_system_taxid: 562. Synthetic: yes. Other_details: synthetic peptide

NMR struc:

20 models

Authors:

M.Hobeika,C.Brockmann,F.Gruessing,D.Neuhaus,G.Divita,M.Stewart, C.Dargemont

Key ref:

M.Hobeika et al. (2009). Structural Requirements for the Ubiquitin-associated Domain of the mRNA Export Factor Mex67 to Bind Its Specific Targets, the Transcription Elongation THO Complex Component Hpr1 and Nucleoporin FXFG Repeats. J Biol Chem, 284, 17575-17583. PubMed id: 19401465 DOI: 10.1074/jbc.M109.004374

Date:

06-Apr-09 Release date: 12-May-09

Protein chain

Q99257  (MEX67_YEAST) -  mRNA export factor MEX67 from Saccharomyces cerevisiae (strain ATCC 204508 / S288c)

Seq: 599 a.a.

Struc: 57 a.a.

Enzyme reactions

• Enzyme class: E.C.?

DOI no: 10.1074/jbc.M109.004374

J Biol Chem 284:17575-17583 (2009)

PubMed id: 19401465

Structural Requirements for the Ubiquitin-associated Domain of the mRNA Export Factor Mex67 to Bind Its Specific Targets, the Transcription Elongation THO Complex Component Hpr1 and Nucleoporin FXFG Repeats.

M.Hobeika, C.Brockmann, F.Gruessing, D.Neuhaus, G.Divita, M.Stewart, C.Dargemont.

ABSTRACT

The ubiquitin-associated (UBA) domain of the principal Saccharomyces cerevisiae mRNA nuclear export factor, Mex67, can bind both nuclear pore protein (nucleoporin) FG repeats and Hpr1, a component of the TREX.THO complex that functions to link transcription and export. Using fluorescence resonance energy transfer-based assays, we show here that Hpr1 and the FG repeats interact with overlapping binding sites on the Mex67 UBA domain. We present the solution structure of the Mex67 UBA domain (UBA-Mex67) complexed with a FXFG nucleoporin peptide and define residues engaged in the interaction and those involved in the FXFG-induced conformational change. We show by NMR titration that the binding of Hpr1 produces analogous changes in chemical shifts in similar regions of the UBA domain. Together the data presented here indicate that both Hpr1 and FXFG nucleoporins may bind in a similar way to the UBA-Mex67 domain. However, whereas binding of Hpr1 allows UBA-Mex67 to interact with tetra-ubiquitin, the complex between UBA-Mex67 and FXFG is unable to bind mono- or tetra-ubiquitin, suggesting that both substrate binding and also the nature of the substrate may influence the affinity of the UBA-Mex67 domain for ubiquitin.

Selected figure(s)

Figure 1.
Hpr1 and FXFG repeats have comparable affinity for the UBA domain of Mex67.A, binding of FXFG to UBA-Mex67 (400 nm) was followed by measuring tryptophan emission at 340 nm upon excitation at 290 nm. B, binding of FXFG, FF18-Nsp1, and Hpr1-C to UBA-Mex67 was monitored by adding increasing concentrations of FITC-labeled proteins (UBA-Mex67 partners) to dansyl-labeled UBA-Mex67 (400 nm) and measuring the quenching of dansyl fluorescence at 440 nm, upon excitation at 350 nm. The titration curves were fitted according to a quadratic equation using Grafit Software (Erithacus Software Ltd.) (40).

Figure 4.
Chemical shift mapping of UBA-Mex67 residues upon binding of FXFG and Hpr1. Normalized combined chemical shift difference Δδ upon binding of Hpr1-C (A) and the FXFG peptide (B) is mapped onto the structure of the UBA-Mex67 domain. Residues showing a normalized Δδ more than 0.6 upon binding of Hpr1 are shown in dark green, whereas residues that show a normalized Δδ more than 0.3 are shown in light green. Corresponding residues in the FXFG titrations are shown in orange and yellow. The position of the FXFG peptide is shown in blue. C, the same data mapped onto the sequence to enable direct comparison.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2009, 284, 17575-17583) copyright 2009.

Figures were selected by an automated process.