PDBsum entry 1jb4

Protein transport

PDB id: 1jb4

Contents

Protein chains

123 a.a. *

Waters ×183

* Residue conservation analysis

PDB id:

1jb4

Name:

Protein transport

Title:

Crystal structure of ntf2 m102e mutant

Structure:

Nuclear transport factor 2. Chain: a, b. Engineered: yes. Mutation: yes

Source:

Rattus norvegicus. Norway rat. Organism_taxid: 10116. Expressed in: escherichia coli. Expression_system_taxid: 562

Biol. unit:

Dimer (from PQS)

Resolution:

2.23Å R-factor: 0.204 R-free: 0.241

Authors:

C.Chaillan-Huntington,P.J.Butler,J.A.Huntington,D.Akin,C.Feldherr, M.Stewart

Key ref:

C.Chaillan-Huntington et al. (2001). NTF2 monomer-dimer equilibrium. J Mol Biol, 314, 465-477. PubMed id: 11846560 DOI: 10.1006/jmbi.2001.5136

Date:

01-Jun-01 Release date: 13-Mar-02

Protein chains

P61972  (NTF2_RAT) -  Nuclear transport factor 2 from Rattus norvegicus

Seq: 127 a.a.

Struc: 123 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

DOI no: 10.1006/jmbi.2001.5136

J Mol Biol 314:465-477 (2001)

PubMed id: 11846560

NTF2 monomer-dimer equilibrium.

C.Chaillan-Huntington, P.J.Butler, J.A.Huntington, D.Akin, C.Feldherr, M.Stewart.

ABSTRACT

Nuclear transport factor 2 (NTF2) mediates nuclear import of RanGDP, a central component of many nuclear trafficking pathways. NTF2 is a homodimer and each chain has independent binding sites for RanGDP and nuclear pore proteins (nucleoporins) that contain FxFG sequence repeats. We show here that the monomer-dimer dissociation constant for NTF2 obtained by sedimentation equilibrium ultracentrifugation is in the micromolar range, indicating that a substantial proportion of cellular NTF2 may be monomeric. To investigate the functional significance of NTF2 dimerization, we engineered a series of point mutations at the dimerization interface and one of these (M118E) remained monomeric below concentrations of 150 microM. CD spectra and X-ray crystallography showed that M118E-NTF2 preserved the wild-type NTF2 fold, although its thermal stability was 20 deg. C lower than that of the wild-type. M118E-NTF2 bound both RanGDP and FxFG nucleoporins less strongly, suggesting that dissociation of the NTF2 dimer could facilitate RanGDP release and thus nucleotide exchange after it had been transported into the nucleus. Moreover, colloidal gold coated with M118E-NTF2 showed reduced binding to Xenopus oocyte nuclear pores. Overall, our results indicate that dimer formation is important for NTF2 function and give insight into the formation of heterodimers by mRNA export factors such as TAP1 and NXT1 that contain NTF2-homology domains.

Selected figure(s)

Figure 2.
Figure 2. Location of M84, M102 and M118 at the NTF2 dimerization interface. An illustration of the side-chains protruding into the interaction interface between NTF2 chains in the dimer. Methionine residues 84, 102 and 118 are all important components of the hydrophobic centre of the interaction interface. Hydrophobic residues are in grey, acidic residues in red, basic residues in blue and amphipolar residues in white. Also shown are the corresponding side-chains in the putative structures (see Suyama et al.[24]) of TAP1 and NXT1. Note that neither TAP1 nor NXT1 contains the equivalent of His100 in NTF2.

Figure 4.
Figure 4. Putative Glu84-His100 salt-bridge stabilizes the M84E-NTF2 dimer. A portion of the 2F[o] -F[c] electron density map of the crystal structure of M84E-NTF2 showing putative interactions between Glu84 and His100 that could contribute to the stability of the dimerization interface. The A chain is yellow and the B chain is blue. The interaction between chains is mediated by a water molecule.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2001, 314, 465-477) copyright 2001.

Figures were selected by an automated process.