Biophysical Society Thematic Meeting | Ascona, Switzerland

Liposomes, Exosomes, and Virosomes: From Modeling Complex Membrane Processes to Medical Diagnostics and Drug Delivery

Poster Abstracts

53-POS Board 27 Quiet OmpG Nanopores for Small Molecule Detection Raghavendar Reddy Sanganna Gari , Patrick Seelheim.Lukas Tamm. University of Virginia, Charlottesville, VA, USA.

Interest in nanopore technology has been growing due to their unique capabilities in small molecule sensing, measurement of protein folding, and low cost DNA and RNA sequencing. The heptameric α-hemolysin (αHL) nanopore has been used successfully for these applications. However, modifications of αHL pores for optimizing the performance are challenging due to its heptameric nature. The E. coli outer membrane protein OmpG is an excellent alternative to αHL because its pore is formed by a single polypeptide chain. OmpG is a 14-stranded β-barrel protein with seven long extracellular flexible loops. It is known that loop 6 controls the pore access in a pH-dependent manner and its dynamics result in spontaneous gating of the open pore current. This phenomenon eventually limits the applications of OmpG in biosensing. Our goal is to develop quiet OmpG nanopores that are free of spontaneous gating and also insensitive to pH. In this work, we achieved quiet pores by altering the gating loop 6 and residues nearby. Here, we present NMR studies of these quiet pores in lipid micelles to assess their loop dynamics and conformational changes along with electrophysiology measurements. Our engineered OmpG nanopores are approximately 50 times quieter than the wildtype protein and are more robust to pH changes. We further modified the inside of these quiet OmpG derivatives with a copper chelating moiety to facilitate the detection of small molecules such as catecholamines and amino acids.

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