Isolation and pharmacological characterization of AdTx1, a natural peptide displaying specific insurmountable antagonism of the α1A-adrenoceptor

L. Quinton 1 Emmanuelle Girard 2, 3 A. Maiga 4 M. Rekik 5 P. Lluel 5 G. Masuyer 6 M. Larregola 7 Catherine Marquer 4 Justina Ciolek 8 T. Magnin 9 R Wagner 9 Jordi Molgó 2 Robert Thai 10 Carole Fruchart-Gaillard 10 Gilles Mourier 11 Julia Chamot-Rooke 1 André Ménez 12 S. Palea 5 Denis Servent 11 Nicolas Gilles 11
1 DCMR - Laboratoire des mécanismes réactionnels
2 INAF - Institut de Neurobiologie Alfred Fessard
3 NBCM - Laboratoire de neurobiologie cellulaire et moléculaire
4 Centre de recherche du CEA/DSV/iBiTec-S/SIMOPRO
5 Urosphère SAS
6 University of Bath [Bath]
7 SU FM - Sorbonne Université - Faculté de Médecine
8 Nicolaus Copernicus University [Toruń]
9 UMR7175, ESBS, LCI,Dept Recepteurs & Prot Membranaires
10 SIMOPRO - Service d'Ingénierie Moléculaire des Protéines
11 LTMB - Laboratoire de Toxinologie Moléculaire et Biotechnologies
SIMOPRO - Service d'Ingénierie Moléculaire des Protéines : DRF/JOLIOT
12 Département d'Ingénierie et d'Etudes des Protéines
Abstract : BACKGROUND AND PURPOSE: Venoms are a rich source of ligands for ion channels, but very little is known about their capacity to modulate G-protein coupled receptor (GPCR) activity. We developed a strategy to identify novel toxins targeting GPCRs. EXPERIMENTAL APPROACH: We studied the interactions of mamba venom fractions with alpha(1)-adrenoceptors in binding experiments with (3)H-prazosin. The active peptide (AdTx1) was sequenced by Edman degradation and mass spectrometry fragmentation. Its synthetic homologue was pharmacologically characterized by binding experiments using cloned receptors and by functional experiments on rabbit isolated prostatic smooth muscle. KEY RESULTS: AdTx1, a 65 amino-acid peptide stabilized by four disulphide bridges, belongs to the three-finger-fold peptide family. It has subnanomolar affinity (K(i)= 0.35 nM) and high specificity for the human alpha(1A)-adrenoceptor subtype. We showed high selectivity and affinity (K(d)= 0.6 nM) of radio-labelled AdTx1 in direct binding experiments and revealed a slow association constant (k(on)= 6 x 10(6).M(-1).min(-1)) with an unusually stable alpha(1A)-adrenoceptor/AdTx1 complex (t(1/2diss)= 3.6 h). AdTx1 displayed potent insurmountable antagonism of phenylephrine's actions in vitro (rabbit isolated prostatic muscle) at concentrations of 10 to 100 nM. CONCLUSIONS AND IMPLICATIONS: AdTx1 is the most specific and selective peptide inhibitor for the alpha(1A)-adrenoceptor identified to date. It displays insurmountable antagonism, acting as a potent relaxant of smooth muscle. Its peptidic nature can be exploited to develop new tools, as a radio-labelled-AdTx1 or a fluoro-labelled-AdTx1. Identification of AdTx1 thus offers new perspectives for developing new drugs for treating benign prostatic hyperplasia.
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Submitted on : Wednesday, January 30, 2019 - 4:19:54 PM
Last modification on : Monday, April 15, 2019 - 5:10:03 PM

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CEA | SORBONNE-UNIVERSITE | NBCM | X-DCMR | X | JOLIOT | DMTS | X-DEP-CHIM | PARISTECH | CEA-DRF

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L. Quinton, Emmanuelle Girard, A. Maiga, M. Rekik, P. Lluel, et al.. Isolation and pharmacological characterization of AdTx1, a natural peptide displaying specific insurmountable antagonism of the α1A-adrenoceptor. British Journal of Pharmacology, Wiley, 2010, 159 (2), pp.316-325. ⟨10.1111/j.1476-5381.2009.00532.x⟩. ⟨cea-02000545⟩

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