Screening for Influx through Membrane Proteins on the Single-Molecule Level using an Automated and Parallel Lipid Bilayer Platform

Abstract

Planar lipid bilayers can be automatically formed by remotely actuated painting in parallel on Micro-Electrode-Cavity-Arrays (MECA)-Chips. Voltage-clamp recording of ionic current through bacterial channels in bilayers is a powerful technique, yielding information on both the physicochemical characteristics and the physiological role of membrane proteins. Antibiotics penetrate bacterial membranes through general diffusion protein channels, like OmpF in E. coli. The MECA chip approach supports the analysis of translocation events in high resolution by detection of the transient block of ionic currents when a single molecule resides in the pore. Here, reconstituting the porins CarO from A. Baumannii, OmpC and OmpF from E. coli and orthologs in K. pneumoniae/E. cloacae, we resolved the transport of antibiotics and thereby study kinetics of antibiotics permeation through porins. This information can be useful in optimizing the design of new antibacterial drugs effective against resistant bacteria.