ja311080j_si_002.pdf (411 kB)
Nanoscale Mechanism of Molecular Transport through the Nuclear Pore Complex As Studied by Scanning Electrochemical Microscopy
journal contribution
posted on 2013-02-13, 00:00 authored by Jiyeon Kim, Anahita Izadyar, Nikoloz Nioradze, Shigeru AmemiyaThe nuclear pore complex (NPC) is the proteinaceous nanopore
that
solely mediates molecular transport across the nuclear envelope between
the nucleus and cytoplasm of a eukaryotic cell. Small molecules (<40
kDa) diffuse through the large pore of this multiprotein complex.
A passively impermeable macromolecule tagged with a signal peptide
is chaperoned through the nanopore by nuclear transport receptors
(e.g., importins) owing to their interactions with barrier-forming
proteins. Presently, this bimodal transport mechanism is not well
understood and is described by controversial models. Herein, we report
on a dynamic and spatially resolved mechanism for NPC-mediated molecular
transport through nanoscale central and peripheral routes with distinct
permeabilities. Specifically, we develop a nanogap-based approach
of scanning electrochemical microscopy to precisely measure the extremely
high permeability of the nuclear envelope to a small probe molecule,
(ferrocenylmethyl)trimethylammonium. Effective medium theories
indicate that the passive permeability of 5.9 × 10–2 cm/s corresponds to the free diffusion of the probe molecule through
∼22 nanopores with a radius of 24 nm and a length of 35 nm.
Peripheral routes are blocked by wheat germ agglutinin to yield 2-fold
lower permeability for 17 nm-radius central routes. This lectin is
also used in fluorescence assays to find that importins facilitate
the transport of signal-tagged albumin mainly through the 7 nm-thick
peripheral route rather than through the sufficiently large central
route. We propose that this spatial selectivity is regulated by the
conformational changes in barrier-forming proteins that transiently
and locally expand the impermeably thin peripheral route while blocking
the central route.