“After many years of intense electrophysiological recordings we prompted the idea of applying the patch clamp principle to the crystallization of membrane proteins. The biophysical basis for this idea emerged while performing single channel recordings in the cell-attached configuration at high acetylcholine concentrations. To rescue the Tc-nAChR from the desensitized state, we manipulated the resting membrane potential (RMP) in the patched membrane. The principle of arresting the conformational changes of a membrane protein using the RMP has never been conceived in a crystallization experiment. Our central hypothesis is that RMP can be used to restrict the conformation freedom of the membrane protein to facilitate crystallization. This hypothesis builds on strong preliminary data obtained using our first prototype of the RMP@LMx device (U.S. patent 10,155,221), engineered in our laboratory (www.nachrs.org) to crystallize the torpedo californica muscle-type nicotinic acetylcholine receptor (Tc-nAChR). The Tc-nAChR is a prime example of a multimeric membrane complex that is one of the most widely studied ion channels, but its high-resolution structure has remained elusive. We are now using the RMP@LMx V1.5 device to perform Resting Membrane Potential high-throughput crystallization screening (RMP-HTCS) of several nicotinic acetylcholine receptors. A high-resolution structure of the nAChR and its complexes with various nicotinic ligands is of crucial importance for the design of novel agents that target defined nervous system pathologies such as Alzheimer’s disease, schizophrenia, depression, attention deficit hyperactivity disorder, and tobacco addiction”. – JALD
The RMP@LMx U.S patent 10,155,221