An Activity-Based Protein Profiling Probe for the Nicotinic Acetylcholine Receptor
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...
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Published in | Journal of the American Chemical Society Vol. 130; no. 47; pp. 15766 - 15767 |
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Main Authors | , , |
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26.11.2008
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Abstract | 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. |
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AbstractList | 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. |
Author | Tantama, Mathew Lin, Wan-Chen Licht, Stuart |
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BackLink | https://www.ncbi.nlm.nih.gov/pubmed/18975901$$D View this record in MEDLINE/PubMed |
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Notes | ark:/67375/TPS-X6VT0RFV-2 Synthetic, electrophysiology, and biochemical methods; discussion of blockade models and voltage-dependence of blockade; statistical analysis of live-cell labeling; labeling results in the absence of light or BPyneTEA. This material is available free of charge via the Internet at https://pubs.acs.org. istex:4CDCC10474CF6400A031E62B908DE8690CC1DC20 ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 |
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Snippet | Activity-based protein profiling (ABPP) has been used extensively to characterize the physiological functions of enzymes but has not yet been extended to ion... |
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SubjectTerms | Cell Line Chemistry Chemistry, Multidisciplinary Electrophysiology Fluorescent Dyes - chemistry Humans Molecular Structure Photobleaching Physical Sciences Receptors, Nicotinic - chemistry Receptors, Nicotinic - metabolism Science & Technology |
Title | An Activity-Based Protein Profiling Probe for the Nicotinic Acetylcholine Receptor |
URI | http://dx.doi.org/10.1021/ja805868x https://api.istex.fr/ark:/67375/TPS-X6VT0RFV-2/fulltext.pdf http://gateway.webofknowledge.com/gateway/Gateway.cgi?GWVersion=2&SrcApp=Summon&SrcAuth=ProQuest&DestApp=WOS&DestLinkType=FullRecord&UT=000263319600012 https://www.ncbi.nlm.nih.gov/pubmed/18975901 https://search.proquest.com/docview/66692685 |
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