NMR solution structure of Cn12, a novel peptide from the Mexican scorpion Centruroides noxius with a typical β‐toxin sequence but with α‐like physiological activity

Abstract

Cn12 isolated from the venom of the scorpion Centruroides noxius has 67 amino-acid residues, closely packed with four disulfide bridges. Its primary structure and disulfide bridges were determined. Cn12 is not lethal to mammals and arthropods in vivo at doses up to 100 lg per animal. Its 3D structure was determined by proton NMR using 850 distance constraints, 36 / angles derived from 36 coupling constants obtained by two different methods, and 22 hydrogen bonds. The overall structure has a two and half turn a-helix (residues 24-32), three strands of antiparallel b-sheet (residues 2-4, 37-40 and 45-48), and a type II turn (residues 41-44). The amino-acid sequence of Cn12 resembles the b scorpion toxin class, although patch-clamp experiments showed the induction of supplementary slow inactivation of Na + channels in F-11 cells (mouse neuroblastoma N18TG-2 · rat DRG2), which means that it behaves more like an a scorpion toxin. This behaviour prompted us to analyse Na + channel binding sites using information from 112 Na + channel gene clones available in the literature, focusing on the extracytoplasmic loops of the S5-S6 transmembrane segments of domain I and the S3-S4 segments of domain IV, sites considered to be responsible for binding a scorpion toxins.Keywords: Centruroides noxius; NMR structure; patchclamp; scorpion toxin; sodium channel.Scorpion toxins are relatively short peptides with a variable length of amino acids, showing characteristic 3D folding comprised of an a-helix and three segments of antiparallel b-sheet structure, stabilized by several disulfide bridges [1][2][3][4][5]. Their known physiological role is to block or modify ion-channel function, causing impairment of cellular communication, which leads to the depolarization of excitable membranes and might cause death of animals stung by scorpions [6,7].There are several reasons why the molecular basis of toxin specificity and molecular mechanism of action continue to be of scientific interest: (a) to study ion channels, the target molecules of most known scorpion toxins, in order to understand their molecular structure and function, thus learning more about cellular excitability; (b) to understand the toxic effects of scorpion venoms, a prerequisite for the development of more effective and safer antidotes and/or vaccines; (c) to find toxins specific for invertebrate organisms with a view to developing biodegradable drugs for pest control; (d) to discover other possible unknown target molecules for which peptides were evolved in the venom of scorpions. The last of these is not trivial, as the huge variability of these peptides, estimated to be of the order of 100 000 in scorpion venom alone, and of which only about 0.2% have been identified, leaves a wide open field for research [4,5,8]. Several recent articles and reviews have reported on the structural and functional aspects of these peptides [3,[8][9][10][11][12][13][14][15][16][17][18]. Most dealt with scorpion toxins as ion-channels blockers or modifiers of their function. However, the st...

Global Transcriptome Analysis of the Scorpion Centruroides noxius: New Toxin Families and Evolutionary Insights from an Ancestral Scorpion Species

“…In the particular case of C. noxius , the sodium and potassium channel toxins have been described in our research group. They include some mammal-specific (Cn2–4 and Cn6–9) and insect-specific (Cn1, 5, 10–12) NaTxs [34], [35], the slotoxin, noxiustoxin, cobatoxin and Erg-toxins, these belonging to the KTx family [36]–[40].…”

“…3A), several conclusions can be drawn. First, it is interesting to notice that even though the biochemical profile of C. noxius venom shows that only toxin Cn12 clearly belongs to the α subfamily [35], there are other assembled transcripts that are grouped in the α clade (blue branches) together with α-NaTxs from C. sculpturatus, M. martensii, P. granulatus and L. mucronatus . In the case of the β type, only the Cn1, 2, 4 and 10 toxins were obtained with identity values >90%, however, other branches in the topology showed assembled isotigs that are closely related to Cn5 and CngtII from C. noxius , CssIX from C. suffusus suffusus , CeII8 from C. elegans and Hj1a and Hj1b from H. judaicus .…”

Mass Fingerprinting of the Venom and Transcriptome of Venom Gland of Scorpion Centruroides tecomanus

“…S2-letter A). Peptides Ct18 (contig 9), Ct22 (contig 16) and Ct21 (contig 13) showed 90, 88 and 91% identity to Cn12 from the scorpion C. noxius , which is structurally similar to the β-ScTX, but has an α-ScTx effect [65].…”

Differential effects of five ‘classical’ scorpion β-toxins on rNav1.2a and DmNav1 provide clues on species-selectivity

“…Cn12, a peptide from Centruroides noxius was purified in (del Rio-Portilla et al, 2004. This toxin appeared to possess a typical β-toxin sequence but an α-like physiological activity.…”