title: Identification of a novel snake peptide toxin displaying high affinity and antagonist behaviour for the α2-adrenoceptors.
abstract: BACKGROUND AND PURPOSE Muscarinic and adrenergic G protein-coupled receptors (GPCRs) are the targets of rare peptide toxins isolated from snake or cone snail venoms. We used a screen to identify novel toxins from Dendroaspis angusticeps targeting aminergic GPCRs. These toxins may offer new candidates for the development of new tools and drugs. EXPERIMENTAL APPROACH In binding experiments with (3) H-rauwolscine, we studied the interactions of green mamba venom fractions with α(2) -adrenoceptors from rat brain synaptosomes. We isolated, sequenced and chemically synthesized a novel peptide, ρ-Da1b. This peptide was pharmacologically characterized using binding experiments and functional tests on human α(2)-adrenoceptors expressed in mammalian cells. KEY RESULTS ρ-Da1b, a 66-amino acid peptide stabilized by four disulphide bridges, belongs to the three-finger-fold peptide family. Its synthetic homologue inhibited 80% of (3) H-rauwolscine binding to the three α(2)-adrenoceptor subtypes, with an affinity between 14 and 73 nM and Hill slopes close to unity. Functional experiments on α(2A) -adrenoceptor demonstrated that ρ-Da1b is an antagonist, shifting adrenaline activation curves to the right. Schild regression revealed slopes of 0.97 and 0.67 and pA(2) values of 5.93 and 5.32 for yohimbine and ρ-Da1b, respectively. CONCLUSIONS AND IMPLICATIONS ρ-Da1b is the first toxin identified to specifically interact with α(2)-adrenoceptors, extending the list of class A GPCRs sensitive to toxins. Additionally, its affinity and atypical mode of interaction open up the possibility of its use as a new pharmacological tool, in the study of the physiological roles of α(2)-adrenoceptor subtypes.
authors.affiliation: CEA, iBiTec-S, Service d'Ingénierie Moléculaire des Protéines, Gif sur Yvette, France.
authors.forename: Céline
authors.initials: C
authors.lastname: Rouget
authors.forename: Loïc
authors.initials: L
authors.lastname: Quinton
authors.forename: Arhamatoulaye
authors.initials: A
authors.lastname: Maïga
authors.forename: Céline
authors.initials: C
authors.lastname: Gales
authors.forename: Geoffrey
authors.initials: G
authors.lastname: Masuyer
authors.forename: Christian
authors.initials: C
authors.lastname: Malosse
authors.forename: Julia
authors.initials: J
authors.lastname: Chamot-Rooke
authors.forename: Robert
authors.initials: R
authors.lastname: Thai
authors.forename: Gilles
authors.initials: G
authors.lastname: Mourier
authors.forename: Edwin
authors.initials: E
authors.lastname: De Pauw
authors.forename: Nicolas
authors.initials: N
authors.lastname: Gilles
authors.forename: Denis
authors.initials: D
authors.lastname: Servent
mesh_terms: D058669:Adrenergic alpha-2 Receptor Antagonists / Q000378:metabolism* / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive; D019556:COS Cells; D002522:Chlorocebus aethiops; D004546:Elapid Venoms / Q000302:isolation & purification / Q000378:metabolism* / Q000494:pharmacology*; D017815:Elapidae; D006801:Humans; D008969:Molecular Sequence Data; D010455:Peptides / Q000378:metabolism* / Q000494:pharmacology; D011485:Protein Binding / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D018341:Receptors, Adrenergic, alpha-2 / Q000378:metabolism*
chemical_list: D058669:Adrenergic alpha-2 Receptor Antagonists; D004546:Elapid Venoms; D010455:Peptides; D018341:Receptors, Adrenergic, alpha-2