Towards an Artificial Replication of the Nuclear Pore Complex

Abstract

The nuclear pore complex acts as a selective barrier between the nucleus and the cytoplasm of a eukaryotic cell, allowing for selective and active transport of biomolecules across the nuclear envelope. The selectivity of the transport is facilitated by shuttle proteins that are able to bind specifically to cargo proteins, actively guide them through the nuclear envelope, and release them on the other side. In this work, an artificial replication of the nuclear transport using poly(methacrylic acid) (PMAA) and poly(N-(2-hydroxyethyl) acrylamide) (PHEAA) as a shuttle and barrier respectively was developed to allow for selective transport of macromolecules through solid-state nanopores. PHEAA was grafted from gold by atom transfer radical polymerisation (ATRP). In water, hydrated "polymer brushes" have been reported to serve as entropic barriers against proteins and other macromolecules. Intermolecular interaction studies using surface plasmon resonance (SPR) and quartz crystal microbalance dissipation (QCM-D) showed that PMAA was able to interact with PHEAA and insert itself into the brush. Further, the interaction was shown to be controllable by changing the pH. This changed PMAA from a protonated state where it could continuously form and break hydrogen bonds with the PHEAA brush, to a charged state where it could not. Fluorescence imaging showed that PMAA with covalently attached fluorescent cargo could undergo pH dependent diffusion through nanopores covered with PHEAA. At a pH of 7.5 PMAA was charged and would be blocked by the polymer brush. At a pH of 4.0 PMAA was protonated and would diffuse through the nanopore and carry the fluorescence cargo with it to the other side. This demonstrated that the artificial system could mimic the key features of the nuclear pore complex and selectively transport molecular cargo across a solid state nanopore.