A Fatty Acid Induces the Functional Assembly of a Channel Protein into Phospholipid Vesicles

Abstract

Nanometer-sized pores (nanopores) have emerged as a novel single-molecule detection technique to probe biomolecules (protein, DNA, and RNA) electrically and/or optically whilst being threaded into the pore. Since then, naturally-occurring pores known as biological nanopores (membrane proteins and bacteriophage portal proteins) along with synthetically-assembled nanopores known as solid-state nanopores have been employed to study various biomolecular characteristics. A third class of nanopores, known as hybrid nanopores, exploits the robust framework and mechanical stability of SS nanopores with atomically reproducible and tunable characteristics of biological nanopores. Biological nanopores supported by robust synthetic membrane eliminates fragility of lipid bilayers and arbitrary geometry of SS nanopores. Our group recently demonstrated the lipid insertion and stable formation of hydrophilic G20C portal protein transmembrane channels, derived from the G20C thermostable virus. Herein, we demonstrate the improved performance of our system by substituting the lipid bilayer support of the protein by thin SiN x free-standing membrane and making further modifications to chemically fix the protein on the membrane support. Finally, we report the application of our hybrid system in nanopore-based sensing of biomolecules.