Transport receptor occupancy in nuclear pore complex mimics

Transport receptor occupancy in nuclear pore complex mimics

Fragasso, A. (TU Delft BN/Cees Dekker Lab; Kavli institute of nanoscience Delft)
de Vries, Hendrik W. (Rijksuniversiteit Groningen)
Andersson, John (Chalmers University of Technology)
van der Sluis, E.O. (TU Delft BN/Marileen Dogterom Lab; Kavli institute of nanoscience Delft)
van der Giessen, Erik (Rijksuniversiteit Groningen)
Onck, Patrick R. (Rijksuniversiteit Groningen)
Dekker, C. (TU Delft BN/Cees Dekker Lab; Kavli institute of nanoscience Delft)

2022

Nuclear pore complexes (NPCs) regulate all molecular transport between the nucleus and the cytoplasm in eukaryotic cells. Intrinsically disordered Phe-Gly nucleoporins (FG-Nups) line the central conduit of NPCs to impart a selective barrier where large proteins are excluded unless bound to a transport receptor (karyopherin; Kap). Here, we assess “Kap-centric” NPC models, which postulate that Kaps participate in establishing the selective barrier. We combine biomimetic nanopores, formed by tethering Nsp1 to the inner wall of a solid-state nanopore, with coarse-grained modeling to show that yeast Kap95 exhibits two populations in Nsp1-coated pores: one population that is transported across the pore in milliseconds, and a second population that is stably assembled within the FG mesh of the pore. Ionic current measurements show a conductance decrease for increasing Kap concentrations and noise data indicate an increase in rigidity of the FG-mesh. Modeling reveals an accumulation of Kap95 near the pore wall, yielding a conductance decrease. We find that Kaps only mildly affect the conformation of the Nsp1 mesh and that, even at high concentrations, Kaps only bind at most 8% of the FG-motifs in the nanopore, indicating that Kap95 occupancy is limited by steric constraints rather than by depletion of available FG-motifs. Our data provide an alternative explanation of the origin of bimodal NPC binding of Kaps, where a stable population of Kaps binds avidly to the NPC periphery, while fast transport proceeds via a central FG-rich channel through lower affinity interactions between Kaps and the cohesive domains of Nsp1. [Figure not available: see fulltext.]

biomimetics
coarse-grained modeling
intrinsically disordered proteins
karyopherins
molecular dynamics
nanopores
nuclear pore complex
nuclear transport receptors

http://resolver.tudelft.nl/uuid:19a5fba0-34cf-448b-9902-1aaf713e612e

https://doi.org/10.1007/s12274-022-4647-1

1998-0124

Nano Research, 15 (11), 9689-9703

Institutional Repository

journal article

© 2022 A. Fragasso, Hendrik W. de Vries, John Andersson, E.O. van der Sluis, Erik van der Giessen, Patrick R. Onck, C. Dekker