Force Spectroscopy of Single Stranded DNA with Biological Nanopore MspA

Abstract

Biological nanopores are single molecule sensors able to probe individual DNA molecules. In this method, a nanometer scale pore is inserted into a lipid bilayer in an electrolyte solution. Voltage is applied across the bilayer and the ionic current through the pore is measured. DNA nucleotides residing in the pore constriction modulate the resistance of the DNA-pore system. Our group previously introduced Mycobacterium smegmatis porin A (MspA) as a biological nanopore sensor with high signal to noise resolution of DNA nucleotides. In this work, we perform force spectroscopy on individual single-stranded DNA molecules. DNA molecules are attached to NeutrAvidin and driven into MspA until the NeutrAvidin comes to rest on MspA's entrance. Approximately 14 nucleotides span the distance to MspA's constriction. The nucleotides within the pore constriction experience a force proportional to the applied voltage. By varying the voltage applied across the pore, we detect stretching of the DNA with angstrom precision. These experiments validate MspA as a single molecule tool to study mechanical properties of DNA.