Constructing ion channels from water-soluble α-helical barrels

The design of peptides that assemble in membranes to form functional ion channels is challenging. Specifically, hydrophobic interactions must be designed between the peptides and at the peptide-lipid interfaces simultaneously. Here, we take a multi-step approach towards this problem. First, we use rational de novo design to generate water-soluble alpha-helical barrels with polar interiors, and confirm their structures using high-resolution X-ray crystallography. These alpha-helical barrels have water-filled lumens like those of transmembrane channels. Next, we modify the sequences to facilitate their insertion into lipid bilayers. Single-channel electrical recordings and fluorescent imaging of the peptides in membranes show monodisperse, cation-selective channels of unitary conductance. Surprisingly, however, an X-ray structure solved from the lipidic cubic phase for one peptide reveals an alternative state with tightly packed helices and a constricted channel. To reconcile these observations, we perform computational analyses to compare the properties of possible different states of the peptide. The de novo design of functional membrane proteins is a formidable challenge. Now, water-soluble peptides have been designed that assemble into alpha-helical barrels with accessible, polar and hydrated central channels. Insights from these structures have been used to produce stable membrane-spanning, cation-selective channels.