Integrating Membrane Transporter Proteins into Droplet Interface Bilayers

Droplet interface bilayers (DIBs) are an emerging tool within synthetic biology that aims to recreate biological processes in artificial cells. A critical component for the utility of these bilayers is controlled flow between compartments and, notably, uphill transport against a substrate concentration gradient. A versatile method to achieve the desired flow is to exploit the specificity of membrane proteins that regulate the movement of ions and transport of specific metabolic compounds. Methods have been in existence for some time to synthesize proteins within a droplet as well as incorporate membrane proteins into DIBS; however, there have been few reports combining synthesis and DIB incorporation for membrane transporters that demonstrate specific, uphill transport. This chapter presents two methods for the incorporation of a membrane transporter into a simple two-droplet DIB system, with the downhill and uphill transport reaction readily monitored by fluorescence microscopy.