Engineered ion channels as emerging tools for chemical biology.

Over the last 25 years, researchers have developed exogenously expressed, genetically engineered, semi-synthetic, and entirely synthetic ion channels. These structures have sufficient fidelity to serve as unique tools that can reveal information about living organisms. One of the most exciting success stories is optogenetics: the use of light-gated channels to trigger action potentials in specific neurons combined with studies of the response from networks of cells or entire live animals. Despite this breakthrough, the use of molecularly engineered ion channels for studies of biological systems is still in its infancy. Historically, researchers studied ion channels in the context of their own function in single cells or in multicellular signaling and regulation. Only recently have researchers considered ion channels and pore-forming peptides as responsive tools to report on the chemical and physical changes produced by other biochemical processes and reactions. This emerging class of molecular probes has a number of useful characteristics. For instance, these structures can greatly amplify the signal of chemical changes: the binding of one molecule to a ligand-gated ion channel can result in flux of millions of ions across a cell membrane. In addition, gating occurs on sub-microsecond time scales, resulting in fast response times. Moreover, the signal is complementary to existing techniques because the output is ionic current rather than fluorescence or radioactivity. And finally, ion channels are also localized at the membrane of cells where essential processes such as signaling and regulation take place. This Account highlights examples, mostly from our own work, of uses of ion channels and pore-forming peptides such as gramicidin in chemical biology. We discuss various strategies for preparing synthetically tailored ion channels that range from de novo designed synthetic molecules to genetically engineered or simply exogenously expressed or reconstituted wild-type channels. Next we consider aspects of experimental design by comparing various membrane environments or systems that make it possible to quantify the response of ion channels to biochemical processes of interest. We present applications of ion channels to answer questions in chemical biology, and propose potential future developments and applications of these single molecule probes. Finally we discuss the hurdles that impede the routine use of ion channel probes in biochemistry and cell biology laboratories and developments and strategies that could overcome these problems. Optogenetics has facilitated breakthroughs in neuroscience, and these results give a dramatic idea of what may lie ahead for designed ion channels as a functional class of molecular probes. If researchers can improve molecular engineering to increase ion channel versatility and can overcome the barriers to collaborating across disciplines, we conclude that these structures could have tremendous potential as novel tools for chemical biology studies.