{"id":6596,"date":"2023-02-17T07:14:50","date_gmt":"2023-02-17T07:14:50","guid":{"rendered":"https:\/\/beta.nanion.de\/resource_library\/2022-chemical-synthesis-of-a-functional-fluorescent-tagged-%ce%b1-bungarotoxin\/"},"modified":"2023-02-17T07:14:50","modified_gmt":"2023-02-17T07:14:50","slug":"2022-chemical-synthesis-of-a-functional-fluorescent-tagged-%ce%b1-bungarotoxin","status":"publish","type":"resource_library","link":"https:\/\/nanion.com\/ja\/resource_library\/2022-chemical-synthesis-of-a-functional-fluorescent-tagged-%ce%b1-bungarotoxin\/","title":{"rendered":"2022 &#8211; Chemical Synthesis of a Functional Fluorescent-Tagged \u03b1-Bungarotoxin"},"content":{"rendered":"<p>\u03b1-bungarotoxin is a large, 74 amino acid toxin containing five disulphide bridges, initially identified in the venom of Bungarus multicinctus snake. Like most large toxins, chemical synthesis of \u03b1-bungarotoxin is challenging, explaining why all previous reports use purified or recombinant \u03b1-bungarotoxin. However, only chemical synthesis allows easy insertion of non-natural amino acids or new chemical functionalities. Herein, we describe a procedure for the chemical synthesis of a fluorescent-tagged \u03b1-bungarotoxin. The full-length peptide was designed to include an alkyne function at the amino-terminus through the addition of a pentynoic acid linker. Chemical synthesis of \u03b1-bungarotoxin requires hydrazide-based coupling of three peptide fragments in successive steps. After completion of the oxidative folding, an azide-modified Cy5 fluorophore was coupled by click chemistry onto the toxin. Next, we determined the efficacy of the fluorescent-tagged \u03b1-bungarotoxin to block acetylcholine (ACh)-mediated currents in response to muscle nicotinic receptor activation in TE671 cells. Using automated patch-clamp recordings, we demonstrate that fluorescent synthetic \u03b1-bungarotoxin has the expected nanomolar affinity for the nicotinic receptor. The blocking effect of fluorescent \u03b1-bungarotoxin could be displaced by incubation with a 20-mer peptide mimicking the \u03b1-bungarotoxin binding site. In addition, TE671 cells could be labelled with fluorescent toxin, as witnessed by confocal microscopy, and this labelling was partially displaced by the 20-mer competitive peptide. We thus demonstrate that synthetic fluorescent-tagged \u03b1-bungarotoxin preserves excellent properties for binding onto muscle nicotinic receptors.<\/p>\n","protected":false},"featured_media":0,"template":"","products_tags":[42],"resource_type":[75],"application_areas":[50],"therapeutic_areas":[],"target_categories":[337],"target_families":[360],"targets":[193],"resource_keyword":[],"class_list":["post-6596","resource_library","type-resource_library","status-publish","hentry","products_tags-syncropatch-384","resource_type-publication","application_areas-ion-channels","target_categories-ligand-gated-ion-channels","target_families-nicotinic-acetylcholine-receptors","targets-achr-1-1---"],"acf":[],"_links":{"self":[{"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/resource_library\/6596","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/resource_library"}],"about":[{"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/types\/resource_library"}],"wp:attachment":[{"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/media?parent=6596"}],"wp:term":[{"taxonomy":"products_tags","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/products_tags?post=6596"},{"taxonomy":"resource_type","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/resource_type?post=6596"},{"taxonomy":"application_areas","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/application_areas?post=6596"},{"taxonomy":"therapeutic_areas","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/therapeutic_areas?post=6596"},{"taxonomy":"target_categories","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/target_categories?post=6596"},{"taxonomy":"target_families","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/target_families?post=6596"},{"taxonomy":"targets","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/targets?post=6596"},{"taxonomy":"resource_keyword","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/resource_keyword?post=6596"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}