| Autor: |
Utkin, Yuri N., Krivoshein, Arcadius V., Davydov, Vasily L., Kasheverov, Igor E., Franke, Peter, Maslennikov, Innokenty V., Arseniev, Alexander S., Hucho, Ferdinand, Tsetlin, Victor I. |
| Předmět: |
|
| Zdroj: |
European Journal of Biochemistry; Apr98 Part 1, Vol. 253 Issue 1, p229-235, 7p, 1 Diagram, 3 Charts, 8 Graphs |
| Abstrakt: |
Different photoactivatable derivatives of toxin 3 (CTX) Naja naja siamensis were obtained after CTX reaction with N-hydroxysuccinimide esters of p-azidobenzoic, p-azidotetraflourobenzoic, p-benzoylbenzoic and p-[3-(trifluoromethyl)-3H-diazirin-3-yl]benzoic acids. The ion-exchange HPLC profiles for the reaction products were very similar in four cases, with one predominant peak corresponding to the derivative containing the label at Lys23. After [125I]iodination, CTX photoactivatable derivatives were cross-linked to the nicotinic acetylcholine receptor from Torpedo californica under optimized conditions. The highest cross-linking yield (up to 16 % of the bound toxin) was observed for azidobenzoyl-Lys23-CTX. Different receptor subunits were found to be labelled depending on the nature of the photoactivatable group : the azido derivatives labelled the γ and δ subunits, benzoylbenzoyl derivative labelled the α and δ subunits, while p-[3-(trifluoromethyl)-3H-diazirin-3-yl]benzoyl derivative reacted with α, γ and δ subunits. The cross-linking experiments in the presence of varying concentrations of (+)-tubocurarine demonstrated that the Lys23-attached diazirinyl group contacts the δ and α subunits in one ligand-binding site, whereas at the other site, for another CTX molecule, the contacts of the Lys23-diazirinyl are with γ and α subunits. This means that the central loop in the two CTX molecules binds at the α/γ and α/δ interfaces. Calculation of the sterically possible displacement of diazirinyl nitrogen, basing on the known X-ray structure of CTX, showed that this value does not exceed 13 Å. The results obtained favor the disposition of the ligand-binding sites at the subunit interfaces, with the distance between α and δ, or α and γ subunits at these sites being not more than 13 Å. [ABSTRACT FROM AUTHOR] |
| Databáze: |
Complementary Index |
| Externí odkaz: |
|
