Autor: E. A. Azeeva, Elena V. Kryukova, Yu. N. Utkin, A. S. Korotina, A. F. Shevalier, V. I. Tsetlin
Rok vydání: 2003
Předmět:
Zdroj: Russian Journal of Bioorganic Chemistry. 29:358-363
ISSN: 1068-1620
Popis: A sensitive nonradioactive method for detection of substances interacting with the neuronal α7-type nicotinic acetylcholine receptor (AChR) was proposed. The method uses biotinylated α-cobratoxin (Bt-CTX) and is based on the ability of the N-terminal ligand-binding extracellular domain (LBED) of AChR to interact with α-cobratoxin (CTX) as does the whole receptor. LBED produced by heterologous expression of a gene fragment of the α7 subunit of AChR from the rat brain in Escherichia coli cells was sorbed in wells of a 96-well plate and incubated with Bt-CTX. The specifically bound Bt-CTX was determined by staining with streptavidin–peroxidase complex. The ability of other compounds to interact with α7-AChR was checked according to the degree with which they inhibit the Bt-CTX binding to LBED. Nicotine, carbamylcholine, d-tubocurarin, anabaseine, conotoxin ImI, and neurotoxin II were used as model compounds. The sensitivity of this method was comparable with that of the radioligand method (up to 10 pmol).
Databáze: OpenAIRE
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