Nanopore Sequencing: Optimizing a Stable Nanopore Platform

Nanopore sequencing is an emerging technology that can provide highly efficient and cost effective DNA sequence analysis compared to conventional sequencing methods. The protein-based nanopore is accommodated in a long-lasting polymer membrane separating two electrolytic solutions, thereby allowing DNA to be electrophoretically passed through the nanopore under an applied voltage. In order to have a successful nanopore sequencer, it’s essential to obtain stability in both the polymer membrane platform and the inserted protein pore. Upon insertion, the protein pore can potentially adopt several conformational states, disrupting the ability to act as a viable sensing device. Protein conformational shape and function rely heavily on their host bilayer and its associated properties such as thickness, curvature, and elastic moduli. This study focuses on the polymer membrane platform and its hydrophobic interactions with the protein nanopore, with the intent of quantitatively understanding the energetics and forces exerted by the polymer on the inserted protein.  We utilize gramicidin A, a 15-amino acid long polypeptide, as a molecular force probe to gain further insight into the forces exerted on the protein by the polymer’s hydrophobic region. A lipid membrane control study is conducted to verify gramicidin A insertion is viable, given all previous scientific data has used gramicidin A to probe solely lipid membranes. Once the membrane forces and their effects on the protein’s conformational states can be quantitatively understood, we can tailor our polymer for the most stable, noise free protein pore insertion.