An Activity-Based Protein Profiling Probe for the Nicotinic Acetylcholine Receptor

• Published in: Journal of the American Chemical Society Vol. 130; no. 47; pp. 15766 - 15767
• Main Authors: Tantama, Mathew, Lin, Wan-Chen, Licht, Stuart
• Format: Journal Article
• Language: English
• Published: WASHINGTON American Chemical Society 26.11.2008; Amer Chemical Soc
• Subjects: Cell Line; Chemistry; Chemistry, Multidisciplinary; Electrophysiology; Fluorescent Dyes - chemistry; Humans; Molecular Structure; Photobleaching; Physical Sciences; Receptors, Nicotinic - chemistry; Receptors, Nicotinic - metabolism; Science & Technology; ALPHA-SUBUNIT; CHANNEL; DESENSITIZATION; BLOCK; COPPER(I)-CATALYZED AZIDE-ALKYNE; AGONIST BINDING; PROTEOMICS
• ISSN: 0002-7863; 1520-5126
• DOI: 10.1021/ja805868x

Activity-based protein profiling (ABPP) has been used extensively to characterize the physiological functions of enzymes but has not yet been extended to ion channels. We have synthesized a state-dependent photoaffinity probe for the nicotinic acetylcholine receptor (nAChR) as a proof of concept for the development of ion channel directed ABPP probes. The candidate probe BPyneTEA comprises an nAChR binding moiety, a benzophenone moiety for photolabeling, and an alkyne moiety for biotinylation via “click chemistry”. Single-molecule current measurements show that BPyneTEA blocks both the closed and open (i.e., nondesensitized) conformations of the nAChR with similar kinetics. In living cells, BPyneTEA photolabels the closed state selectively over the inactive desensitized state. BPyneTEA thus shows promise as a probe for nondesensitized nAChRs and may be useful in studying the molecular physiology of desensitization. The structure and reactivity of ion channel pores in general suggest that they will be a broadly useful target for ABPP probes.