Droplet interface bilayer (DIB) as a model for the study of water and ion transport in membrane proteins and pores

Abstract

Droplet interface bilayers (DIBs) are self-assembled lipid bilayers formed when connecting two water droplets enclosed by a lipid monolayer in a hydrophobic environment. Since the development of the technique in 2006, DIBs have become one of the most promising platforms for constructing artificial tissues. However, mimicking natural cell capabilities depends on incorporating functional molecules such as membrane proteins into the network. So far, only a few pores have been successfully incorporated to create functional DIB tissues, and interactions between DIBs and membrane proteins are not yet fully understood. Another difficulty in designing DIB tissues is that they are formed by passive water transport across the lipid bilayer, which has low permeability, and consequently, it may take hours for small tissues to respond to stimuli. For these reasons, this work aimed to implement the tools for studying membrane proteins and pores of DIBs in Kumar’s Lab, including water and ion transporters. OmpF and Gramicidin were employed as model membrane proteins to validate the methods for measuring capacitance, conductance, and water permeability in DIBs. By setting up an electrophysiology rig, we measured capacitance and conductance by applying a known voltage stimulus and analyzing the current response of the two-droplet DIB system. The results agreed with the theoretical behavior and the results from previous authors. The water permeability of DPhPC was determined by microscopic image analysis of the DIB pair, obtaining an average of 30μm/s. Similar values were observed for DIBs containing gramicidin due most likely to a low density of channels obtained. Although there is still room for improvement, these results accomplish our primary goal, validating the platform for future studies of integral membrane proteins and pore-forming molecules.