Unpublished conference/Abstract (Scientific congresses and symposiums)
New methodology to detect toxin-nAChRs binding by MALDI-TOF mass spectrometry
Echterbille, Julien; Gilles, Nicolas; Araoz, Romulo et al.
2016Third NVMS-BSMS Conference on Mass Spectrometry
 

Files


Full Text
160316-Echterbille Rolduc-V1.pptx
Publisher postprint (1.32 MB)
Request a copy

All documents in ORBi are protected by a user license.

Send to



Details



Keywords :
nAChRs; Toxins; MALDI-TOF
Abstract :
[en] More than 50 thousands of venomous species are currently indexed in the world. Each of their venoms is composed of 200 to 1000 different toxins which potentially exhibit a high selectivity for membrane receptors such as G-protein coupled receptors or ion channels such nicotinic acetylcholine receptors (nAChRs). The latter have been a target for drug discovery efforts, primarily for central nervous system indications. Therefore, it appears of prime interest to design new pharmacological tools and potentially discover future drugs targeting this kind of ion channels. In 2015, our group published a new mass spectrometry-based methodology to screen peptide ligands for G protein coupled receptors1. The proof of concept of this methodology was built by studying the binding of [Arg8]-vasopressin (AVP) on type 2-vasopressin receptor (V2). We extended this methodology to another system ligand-receptor. As all Conus species venoms investigated so far contain at least one toxin antagonizing nAChRs: the alpha-conotoxins. Therefore, the ligand-receptor model couple that has been chosen is nAChRs-alpha-conotoxins. Experimentally, fragments of cellular membranes over-expressing nAChRs were incubated with Bovine Serum Albumine (BSA) tryptic digest (~100 peptide toxins) doped by a small amount of Alpha-conotoxins. After 2 hours incubation, free and bound fractions were purified with a combination of centrifugation and micro column purifications. Samples were finally analyzed with a MALDI-TOF/TOF mass spectrometer. By comparison of the intensity of Alpha-conotoxins in the free and in the bound fractions, we clearly detect an enrichment of nAChRs ligand in the latter. In order to transpose the methodology to natural mixture, we applied the workflow to crude conus venoms. We incubated membranes over-expressing nAChRs with Conus textile venom which is known to possess at least 5 different alpha-conotoxins. Thanks to our approach, we were able to detect an enrichment of these known ligands in the bound fraction. In order to validate the potential of our approach, the next step of this work will be the incubation of a Conus venom for which no alpha-conotoxins have been described.
Disciplines :
Chemistry
Author, co-author :
Echterbille, Julien ;  Université de Liège > Département de chimie (sciences) > Laboratoire de spectrométrie de masse (L.S.M.)
Gilles, Nicolas;  Commissariat à l'Energie Atomique (Saclay) - CEA > DSV/iBiTec-S/SIMOPRO
Araoz, Romulo;  Centre de Recherche CNRS de Gif-sur-Yvette > Institut Fédératif de Neurobiologie Alfred Fessard FR2118 > Laboratoire de Neurobiologie et Développement UPR 3294
De Pauw, Edwin  ;  Université de Liège > Département de chimie (sciences) > Laboratoire de spectrométrie de masse (L.S.M.)
Quinton, Loïc  ;  Université de Liège > Département de chimie (sciences) > Chimie biologique
Language :
English
Title :
New methodology to detect toxin-nAChRs binding by MALDI-TOF mass spectrometry
Alternative titles :
[en] Nouvelle méthodologie pour la détection de la liaison nAChRs-toxine animale par Spectrométrie de Masse MALDI-TOF
Publication date :
18 April 2016
Event name :
Third NVMS-BSMS Conference on Mass Spectrometry
Event organizer :
BSMS et NVMS
Event place :
Rolduc, Netherlands
Event date :
du 17 avril 2016 au 19 avril 2016
Audience :
International
Available on ORBi :
since 27 August 2016

Statistics


Number of views
126 (6 by ULiège)
Number of downloads
0 (0 by ULiège)

Bibliography


Similar publications



Contact ORBi