1qg2
From Proteopedia
|
CANINE GDP-RAN R76E MUTANT
Overview
Nuclear protein import requires a precisely choreographed series of, interactions between nuclear pore components and soluble factors such as, importin-beta, Ran, and nuclear transport factor 2 (NTF2). We used the, crystal structure of the GDPRan-NTF2 complex to design mutants in the, switch II loop of Ran to probe the contribution of Lys71, Phe72 and Arg76, to this interaction. X-ray crystallography showed that the F72Y, F72W and, R76E mutations did not introduce major structural changes into the mutant, Ran. The GDP-bound form of the switch II mutants showed no detectable, binding to NTF2, providing direct evidence that salt bridges involving, Lys71 and Arg76 and burying Phe72 are all crucial for the interaction, between Ran and NTF2. Nuclear protein accumulation in, digitonin-permeabilzed cells was impaired with Ran mutants deficient in, NTF2 binding, confirming that the NTF2-Ran interaction is required for, efficient transport. We used mutants of the yeast Ran homologue Gsp1p to, investigate the effect of the F72Y and R76E mutations in vivo. Although, neither mutant was viable when integrated into the genome as a single, copy, yeast mildly overexpressing the Gsp1p mutant corresponding Ran F72Y, on a centromeric plasmid were viable, confirming that this mutant retained, the essential properties of wild-type Ran. However, yeast expressing the, Gsp1p mutant corresponding to R76E to comparable levels were not viable, although strains overexpressing the mutant to higher levels using an, episomal 2micrometers plasmid were viable, indicating that the R76E, mutation may also have interfered with other interactions made by Gsp1p.
About this Structure
1QG2 is a Single protein structure of sequence from Canis lupus familiaris with MG and GDP as ligands. Full crystallographic information is available from OCA.
Reference
Engineered mutants in the switch II loop of Ran define the contribution made by key residues to the interaction with nuclear transport factor 2 (NTF2) and the role of this interaction in nuclear protein import., Kent HM, Moore MS, Quimby BB, Baker AM, McCoy AJ, Murphy GA, Corbett AH, Stewart M, J Mol Biol. 1999 Jun 11;289(3):565-77. PMID:10356329
Page seeded by OCA on Wed Nov 21 00:39:31 2007
