Ran in Nucleocytoplasmic Transport - Ras Superfamily Small G Proteins: Biology and Mechanisms 2 - page 115

RanGTP binding to karyopherins is able to mediate a change in the helicoidal
pitch by interacting with generally two or three sites dispersed along the molecule
so that the karyopherin tends to be wrapped around RanGTP. Generally all
karyopherins bind RanGTP at a site near their N-terminus and which involves
residues from the first three HEAT repeats of the karyopherin helicoid (Vetter
et al. 1999a ; Chook and Blobel 1999 ; Matsuura and Stewart 2005 ; Lee et al. 2005 ;
Cook et al. 2009 ; Monecke et al. 2009 ; Dong et al. 2009a ; Okada et al. 2009 )
interacting primarily with the switch II loop. The other areas of interaction are not
so well conserved and tend to vary somewhat between different karyopherins. In the
yeast Cse1:Kap60:RanGTP complex, for example, the switch I loop interacts with
both the N-terminal region of Cse1 as well as a flexible loop emanating from near
its C-terminus, while residues centered on Ran Lys152 also interact with the
importin-

α analogue, Kap60 in the complex (Matsuura and Stewart 2004 ). In the

importin- :RanGTP complex, in addition to the interaction with HEAT repeats

1-4 at the importin-

β

N-terminus, basic residues (Lys 134, Lys139, Arg140, and
Lys141) on the opposite face of RanGTP bind at HEAT repeats 7 and 8 and a third
site, centered on Lys152 (analogous to the second Ran-binding site on Cse1), binds
to HEAT repeats 12-15. RanGTP binding at these three sites bends the importin-

β

β
into a more rigid, compact conformation in which the distortion of the helicoid
disrupts the conformation of the extensive IBB-binding region on its inner, con-
cave, surface and thereby facilitates the release of importin- (Lee et al. 2005 ).

RanGTP binding also induces a considerable change in the conformation of
Exportin-t (Los1 in yeast) that results in its somewhat open conformation in
solution becoming more compact and bringing its N- and C-termini closer together
to form a binding site for tRNAs (Cook et al. 2009 ). This mode of binding explains
how Xpo-t can recognize all mature tRNAs in the cell and yet distinguish them from
those that have not been properly processed, thus coupling tRNA export to quality
control (Cook et al. 2009 ). An analogous conformational-change linked molecular
recognition was also seen in the crystal structure of pre-miRNA complexed with
Exportin-5 and RanGTP (Okada et al. 2009 ). This crystal structure shows that
Exportin-5:RanGTP recognizes the 2-nucleotide 3 0 overhang and the double-
stranded stem of the pre-miRNA, shielding it from degradation, whereas a tunnel-
like structure of Exportin-5 interacts strongly with the 2-nucleotide 3 0 overhang
through H-bonds and ionic interactions. Interestingly, RNA recognition in this
complex does not depend on RNA sequence, implying that Exportin-5:RanGTP
can recognize a variety of pre-miRNAs (Okada et al. 2009 ). However, RanGTP
binding appears to cause a much smaller structural alteration in the helicoid in some
other karyopherins, including transportin and CRM1 (Xpo1 in yeast). For CRM1
binding either NES-containing cargoes or the snurportin adaptor is cooperative with
RanGTP binding and a major determinant in modulating the NES-binding site
appears to be the position adopted by a long

α

-helical extension located at the
CRM1 C-terminus that, in the presence of RanGTP, is placed so that a group of
charged residues at its C-terminus is located immediately below the NES-binding
site and so is ideally placed to influence the affinity of CRM1 for NESs (Dong
et al. 2009a , b ; Monecke et al. 2009 ; Fox et al. 2011 ). Comparison with the

α