Nanopores enable the characterization of single biomolecules; to interrogate these molecules without artifacts from wall interactions, nanopore coatings are required.

In the context of sensing and characterizing single proteins with synthetic nanopores, lipid bilayer coatings provide at least four benefits: first, they minimize unwanted protein adhesion to the pore walls by exposing a zwitterionic, fluid surface. Second, they can slow down protein translocation and rotation by the opportunity to tether proteins with a lipid anchor to the fluid bilayer coating. Third, they provide the possibility to impart analyte specificity by including lipid anchors with a specific receptor or ligand in the coating. Fourth, they offer a method for tuning nanopore diameters by choice of the length of the lipid’s acyl chains. The work presented here compares four properties of various lipid compositions with regard to their suitability as nanopore coatings for protein sensing experiments: (1) electrical noise during current recordings through solid-state nanopores before and after lipid coating, (2) long-term stability of the recorded current baseline and, by inference, of the coating, (3) viscosity of the coating as quantified by the lateral diffusion coefficient of lipids in the coating, and (4) the success rate of generating a suitable coating for quantitative nanopore-based resistive pulse recordings. We surveyed lipid coatings prepared from bolaamphiphilic, monolayer-forming lipids inspired by extremophile archaea and compared them to typical bilayer-forming phosphatidylcholine lipids containing various fractions of curvature-inducing lipids or cholesterol. We found that coatings from archaea-inspired lipids provide several advantages compared to conventional phospholipids; the stable, low noise baseline qualities and high viscosity make these membranes especially suitable for analysis that estimates physical protein parameters such as the net charge of proteins as they enable translocation events with sufficiently long duration to time-resolve dwell time distributions completely. The work presented here reveals that the ease or difficulty of coating a nanopore with lipid membranes did not depend significantly on the composition of the lipid mixture, but rather on the geometry and surface chemistry of the nanopore in the solid state substrate. In particular, annealing substrates containing the nanopore increased the success rate of generating stable lipid coatings.

Nano- and micrometer-sized compartments composed of synthetic polymers are designed to mimic spatial and temporal divisions found in nature. Self-assembly of polymers into compartments such as polymersomes, giant unilamellar vesicles (GUVs), layer-by-layer (LbL) capsules, capsosomes, or polyion complex vesicles (PICsomes) allows for the separation of defined environments from the exterior. These compartments can be further engineered through the incorporation of (bio)molecules within the lumen or into the membrane, while the membrane can be decorated with functional moieties to produce catalytic compartments with defined structures and functions. Nanometer-sized compartments are used for imaging, theranostic, and therapeutic applications as a more mechanically stable alternative to liposomes, and through the encapsulation of catalytic molecules, i.e., enzymes, catalytic compartments can localize and act in vivo. On the micrometer scale, such biohybrid systems are used to encapsulate model proteins and form multicompartmentalized structures through the combination of multiple compartments, reaching closer to the creation of artificial organelles and cells. Significant progress in therapeutic applications and modeling strategies has been achieved through both the creation of polymers with tailored properties and functionalizations and novel techniques for their assembly.