Cell-Free Protein Synthesis Goes Electric—Measuring Transmembrane Ion Channel Activity Using Bioelectronic Circuits

The sensor elements of a biological cell are its transmembrane proteins (TMP). Housed in a lipid bilayer, these species carry out a variety of functions that let the cell interact and respond to its environment. Harnessing these functions for biosensing purposes that can lead to new diagnostics, drug screening platforms, and pathogen detection tools are just a few of the promising applications possible if TMP could be integrated with sensing platforms that have an easy readout. However, this class of proteins remains challenging to study. Progress has been hindered by the difficulty in synthesizing, isolating, and incorporating them and their membrane scaffolds into a sensing platform. Cell-free protein synthesis (CFPS) is an emerging technique for synthesizing TMPs directly into a membrane environment by using reconstituted components of the cellular transcription and translation machinery in vitro, bypassing purification and reconstitution into a membrane. Our recent goals have been to leverage this approach to integrate TMPs into supported lipid bilayers coating conducting polymer electronic circuits. Specifically, PEDOT:PSS electrode arrays, which are both transparent and electrically conductive, can be coated with lipid membranes and enable the assessment of protein incorporation and activity by optical and electrical means. We use fluorescence microscopy and electrochemical impedance spectroscopy (EIS) to characterize the synthesized TMPs. In this talk, I will share our success in incorporating several TMP ion channels into biomembrane bioelectronic devices. By dramatically simplifying TMP incorporation into a platform capable of sensing their biological function, label-free, we envision a future where large libraries of protein variants can be easily synthesized and assayed using bioelectronic circuits. This kind of scale up is necessary to mimic and achieve the systems-level sensing used in nature.