Quantifying Permeation of Small Charged Molecules across Channels: Electrophysiology in Small Volumes

Abstract

A major bottleneck in the development of small-molecule antibiotics is to achieve good permeability across the outer membrane in Gram-negative bacteria. Optimization with respect to permeability surprisingly lacks appropriate methods. Recently, we proposed to use the diffusion potential for charged molecules created by their difference in electrophoretic mobility while crossing the outer membrane channel under a concentration gradient. The latter provides semiquantitative values, but the current available setups require large volumes and thus exclude several classes of molecules. Here we propose a simple approach of capturing proteoliposomes at the aperture of glass surface (planar aperture or conical glass capillary) by decreasing the necessary volume below 50 μL. We measured the transport of two charged molecules sulbactam and ceftazidime across the two major porins in Escherichia coli. Both molecules were observed to permeate through these porins, with sulbactam having a higher permeability.