Novel alpha-conotoxins from Conus spurius and the alpha-conotoxin EI share high-affinity potentiation and low-affinity inhibition of nicotinic acetylcholine receptors.
alpha-Conotoxins from marine snails are known to be selective and potent competitive antagonists of nicotinic acetylcholine receptors. Here we describe the purification, structural features and activity of two novel toxins, SrIA and SrIB, isolated from Conus spurius collected in the Yucatan Channel, Mexico. As determined by direct amino acid and cDNA nucleotide sequencing, the toxins are peptides containing 18 amino acid residues with the typical 4/7-type framework but with completely novel sequences. Therefore, their actions (and that of a synthetic analog, [gamma15E]SrIB) were compared to those exerted by the alpha4/7-conotoxin EI from Conus ermineus, used as a control. Their target specificity was evaluated by the patch-clamp technique in mammalian cells expressing alpha(1)beta(1)gammadelta, alpha(4)beta(2) and alpha(3)beta(4) nicotinic acetylcholine receptors. At high concentrations (10 microm), the peptides SrIA, SrIB and [gamma15E]SrIB showed weak blocking effects only on alpha(4)beta(2) and alpha(1)beta(1)gammadelta subtypes, but EI also strongly blocked alpha(3)beta(4) receptors. In contrast to this blocking effect, the new peptides and EI showed a remarkable potentiation of alpha(1)beta(1)gammadelta and alpha(4)beta(2) nicotinic acetylcholine receptors if briefly (2-15 s) applied at concentrations several orders of magnitude lower (EC(50), 1.78 and 0.37 nm, respectively). These results suggest not only that the novel alpha-conotoxins and EI can operate as nicotinic acetylcholine receptor inhibitors, but also that they bind both alpha(1)beta(1)gammadelta and alpha(4)beta(2) nicotinic acetylcholine receptors with very high affinity and increase their intrinsic cholinergic response. Their unique properties make them excellent tools for studying the toxin-receptor interaction, as well as models with which to design highly specific therapeutic drugs.
López-Vera
Estuardo
E
Laboratorio de Neurofarmacología Marina, Departamento de Neurobiología Celular y Molecular, Instituto de Neurobiología, Universidad Nacional Autónoma de México, Campus Juriquilla, Queretaro, México.
Aguilar
Manuel B
MB
Schiavon
Emanuele
E
Marinzi
Chiara
C
Ortiz
Ernesto
E
Restano Cassulini
Rita
R
Batista
Cesar V F
CV
Possani
Lourival D
LD
Heimer de la Cotera
Edgar P
EP
Peri
Francesco
F
Becerril
Baltazar
B
Wanke
Enzo
E
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D020916:Conotoxins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism* / Q000494:pharmacology*; D052078:Conus Snail / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004220:Disulfides / Q000737:chemistry / Q000378:metabolism; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D008970:Molecular Weight; D018733:Nicotinic Antagonists / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism* / Q000494:pharmacology*; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D011978:Receptors, Nicotinic / Q000378:metabolism*; D012680:Sensitivity and Specificity
D020916:Conotoxins; D004220:Disulfides; D018733:Nicotinic Antagonists; D010455:Peptides; D011978:Receptors, Nicotinic