title: Karyopherin-centric control of nuclear pores based on molecular occupancy and kinetic analysis of multivalent binding with FG nucleoporins.
abstract: Intrinsically disordered Phe-Gly nucleoporins (FG Nups) within nuclear pore complexes exert multivalent interactions with transport receptors (Karyopherins (Kaps)) that orchestrate nucleocytoplasmic transport. Current FG-centric views reason that selective Kap translocation is promoted by alterations in the barrier-like FG Nup conformations. However, the strong binding of Kaps with the FG Nups due to avidity contradicts rapid Kap translocation in vivo. Here, using surface plasmon resonance, we innovate a means to correlate in situ mechanistic (molecular occupancy and conformational changes) with equilibrium (binding affinity) and kinetic (multivalent binding kinetics) aspects of Karyopherinβ1 (Kapβ1) binding to four different FG Nups. A general feature of the FxFG domains of Nup214, Nup62, and Nup153 is their capacity to extend and accommodate large numbers of Kapβ1 molecules at physiological Kapβ1 concentrations. A notable exception is the GLFG domain of Nup98, which forms a partially penetrable cohesive layer. Interestingly, we find that a slowly exchanging Kapβ1 phase forms an integral constituent within the FG Nups that coexists with a fast phase, which dominates transport kinetics due to limited binding with the pre-occupied FG Nups at physiological Kapβ1 concentrations. Altogether, our data reveal an emergent Kap-centric barrier mechanism that may underlie mechanistic and kinetic control in the nuclear pore complex.
authors.affiliation: Biozentrum and the Swiss Nanoscience Institute, University of Basel, Basel, Switzerland.
authors.forename: Larisa E
authors.initials: LE
authors.lastname: Kapinos
authors.affiliation: Biozentrum and the Swiss Nanoscience Institute, University of Basel, Basel, Switzerland.
authors.forename: Rafael L
authors.initials: RL
authors.lastname: Schoch
authors.affiliation: Biozentrum and the Swiss Nanoscience Institute, University of Basel, Basel, Switzerland.
authors.forename: Raphael S
authors.initials: RS
authors.lastname: Wagner
authors.affiliation: Biozentrum and the Swiss Nanoscience Institute, University of Basel, Basel, Switzerland.
authors.forename: Kai D
authors.initials: KD
authors.lastname: Schleicher
authors.affiliation: Biozentrum and the Swiss Nanoscience Institute, University of Basel, Basel, Switzerland. Electronic address: roderick.lim@unibas.ch.
authors.forename: Roderick Y H
authors.initials: RY
authors.lastname: Lim
mesh_terms: D000818:Animals; D005998:Glycine / Q000031:analogs & derivatives* / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D022022:Nuclear Pore / Q000378:metabolism*; D028861:Nuclear Pore Complex Proteins / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D020349:Surface Plasmon Resonance; D014982:Xenopus laevis; D028961:beta Karyopherins / Q000378:metabolism*
chemical_list: C424166:KPNB1 protein, human; D028861:Nuclear Pore Complex Proteins; D028961:beta Karyopherins; C008852:2-phenylglycine; D005998:Glycine
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