Single polypeptide detection using a translocon EXP2 nanopore.

Abstract

DNA sequencing using nanopores has already been achieved and commercialized; the next step in advancing nanopore technology is towards protein sequencing. Although trials have been reported for discriminating the 20 amino acids using biological nanopores and short peptide carriers, it remains challenging. The size compatibility between nanopores and peptides is one of the issues to be addressed. Therefore, exploring biological nanopores that are suitable for peptide sensing is key in achieving amino acid sequence determination. Here, we focus on EXP2, the transmembrane protein of a translocon from malaria parasites, and describe its pore-forming properties in the lipid bilayer. EXP2 mainly formed a nanopore with a diameter of 2.5nm assembled from 7 monomers. Using the EXP2 nanopore allowed us to detect poly-L-lysine (PLL) at a single-molecule level. Furthermore, the EXP2 nanopore has sufficient resolution to distinguish the difference in molecular weight between two individual PLL, long PLL (Mw: 30,000-70,000) and short PLL (Mw: 10,000). Our results contribute to the accumulation of information for peptide-detectable nanopores.