A Single-molecule Mycobacterium Smegmatis Porin A Protein Nanopore Sensor for Host-Guest Chemistry

Abstract

The mycobacterium smegmatis porin A (MspA) protein nanopore was prepared by E. coil prokaryotic expression system. The extraction of MspA nanopore using surfactants were carried out under the optimal conditions such as 0.5% extraction agent, 30 min of extraction and at 90 °C. The interactions of MspA nanopore with a variety of cyclodextrins were examined at single-molecule level. The interaction between MspA and Per-6-amino-β-cyclodextrin (am7-β-CD) was stronger and had consistent blocking platform. Its retention time in the MspA nanopore decreased with the increasing voltage, whereas the blocking current was not affected by the voltage. So, am7-β-CD was selected as a non-covalent adapter and lodged within single MspA nanopore to study the single-molecule host-guest chemistry. After adding am7-β-CD to cis side and displaying a first blocked platform, a low concentration of 50 nM for the amantadine hydrochloride could show a second blocked platform and its residence time decreased with the increasing voltage. This new protein nanopore sensor can be used as a single-molecule detector and identifier for small organic molecules, which will greatly broaden the application range of MspA nanopore.