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Correlating ion channel structure and function.

TitleCorrelating ion channel structure and function.
Publication TypeJournal Article
Year of Publication2021
AuthorsSchmidpeter PAM, Nimigean CM
JournalMethods Enzymol
Volume652
Pagination3-30
Date Published2021
ISSN1557-7988
KeywordsCryoelectron Microscopy, Ion Channels, Kinetics, Lipids, Membrane Proteins
Abstract

Recent developments in cryogenic electron microscopy (cryo-EM) led to an exponential increase in high-resolution structures of membrane proteins, and in particular ion channels. However, structures alone can only provide limited information about the workings of these proteins. In order to understand ion channel function and regulation in molecular detail, the obtained structural data need to be correlated to functional states of the same protein. Here, we describe several techniques that can be employed to study ion channel structure and function in vitro and under defined, similar conditions. Lipid nanodiscs provide a native-like environment for membrane proteins and have become a valuable tool in membrane protein structural biology and biophysics. Combined with liposome-based flux assays for the kinetic analysis of ion channel activity as well as electrophysiological recordings, researchers now have access to an array of experimental techniques allowing for detailed structure-function correlations using purified components. Two examples are presented where we put emphasis on the lipid environment and time-resolved techniques together with mutations and protein engineering to interpret structural data obtained from single particle cryo-EM on cyclic nucleotide-gated or Ca2+-gated K+ channels. Furthermore, we provide short protocols for all the assays used in our work so that others can adapt these techniques to their experimental needs. Comprehensive structure-function correlations are essential in order to pharmacologically target channelopathies.

DOI10.1016/bs.mie.2021.02.016
Alternate JournalMethods Enzymol
PubMed ID34059287
Grant ListR01 GM088352 / GM / NIGMS NIH HHS / United States
R01 GM124451 / GM / NIGMS NIH HHS / United States