A substituted cysteine accessibility protocol to probe orthosteric and non-canonical ligand binding to ion channels
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Updated: Nov 4, 2026

Functional Reconstitution and Channel Activity Measurements of Purified Wildtype and Mutant CFTR Protein
Published on: March 9, 2015
Gauri C Kulkarni1, Christian J Peters2
1Department of Anatomy and Cell Biology, University of Illinois Chicago, Chicago, IL 60612, USA.
Abstract:
The Substituted Cysteine Accessibility Method (SCAM) is an approach for probing the biophysics and pharmacology of ion channels by targeting covalent modifications to individual amino acids. Here, we present a protocol to constrain channel stoichiometry and identify asymmetric or non-canonical ligand-binding sites using SCAM followed by electrophysiology in mammalian heterologous expression systems, in the context of heteropentameric nicotinic acetylcholine receptors. This protocol is applicable to examination of multiple channel properties, including ligand binding, gating, pore conductance, and selectivity. For complete information on the use and execution of this protocol, please refer to Kulkarni et al.1.
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