AA sequence: Gly-Cys2-Lys-Gly-Phe-Gly-Asp-Ser-Cys9-Thr-Pro-Gly-Lys-Asn-Glu-Cys16-Cys17-Pro-Asn-Tyr-Ala-Cys22-Ser-Ser-Lys-His-Lys-Trp-Cys29-Lys-Val-Tyr-Leu-NH2
Disulfide bonds: Cys2-Cys17, Cys9-Cys22, and Cys16-Cys29
Length (aa): 33
Formula: C154H228N44O45S6
Molecular Weight: 3608.20 Da
Appearance: White lyophilized solid
Solubility: water and saline buffer
CAS number: Not available
Source: Synthetic
Purity rate: > 97%
Hainantoxin-III
Selective blocker of TTX-S VGSC
Hainantoxin-III (HNTX-III; hainantoxin-3) is a peptide that has been isolated from the venom of the Chinese bird spider Seleconosmia hainana. Hainantoxin-III specifically blocks mammalian neuronal tetrodotoxin-sensitive voltage-gated sodium channels (VGSCs). Hainantoxin III was found inactive on tetrodotoxin-resistant VGSCs and voltage-gated Ca2+channels (both high and low voltage-activated).
Hainantoxin-III strongly depressed the amplitude of rat DRG tetrodotoxin-sensitive Na+ currents with an IC50 value of 1.1 nM. Like Hainantoxin-IV, Hainantoxin-III causes a hyperpolarizing shift of about 10 mV in the voltage midpoint of steady-state Na+ channel inactivation. Similar to Huwentoxin-IV, Hainantoxin-III and Hainantoxin-IV do not affect the activation and inactivation kinetics of Na+ currents.
Hainantoxin-III inhibits Nav1.7 current amplitude without significantly altering the activation and inactivation kinetics. Hainantoxin-III increases the deactivation of the Nav1.7 current after extreme depolarizations. Hainantoxin-III seems to interact with site 4 and to trap the domain II voltage sensor in the closed state. The inhibition of Nav1.7 by hainantoxin-III is reversible upon washing, but no reversibility was observed for Hainantoxin-IV and Huwentoxin-IV. Hainantoxin-III was shown to block Nav1.1, Nav1.2, Nav1.3 and Nav1.7 expressed in HEK293 cells with IC50 values of 1.27 µM, 275 nM, 491 nM and 232 nM, respectively.
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Inhibition of neuronal tetrodotoxin-sensitive Na+ channels by two spider toxins: hainantoxin-III and hainantoxin-IV.
Hainantoxin-III and hainantoxin-IV, isolated from the venom of the Chinese bird spider Seleconosmia hainana, are neurotoxic peptides composed of 33-35 residues with three disulfide bonds. Using whole-cell patch-clamp technique, we investigated their action on ionic channels of adult rat dorsal root ganglion neurons. It was found that the two toxins did not affect Ca2+ channels (both high voltage activated and low voltage activated types) nor tetrodotoxin-resistant voltage-gated Na+ channels (VGSCs). However, hainantoxin-III and hainantoxin-IV strongly depressed the amplitude of tetrodotoxin-sensitive Na+ currents with IC50 values of 1.1 and 44.6 nM, respectively. Both hainantoxin-III (1 nM) and hainantoxin-IV (50 nM) caused a hyperpolarizing shift of about 10 mV in the voltage midpoint of steady-state Na+ channel inactivation, but they showed difference in the reprime kinetics of VGSCs: hainantoxin-III significantly decreased the recovery rate from inactivation at a prepulse potential of -80 mV while hainantoxin-IV did not do. It is interesting to note that similar to huwentoxin-IV, the two hainantoxins did not affect the activation and inactivation kinetics of Na+ currents and at a concentration of 1 microM they completely inhibited the slowing inactivation currents induced by BMK-I (toxin I from the scorpion Buthus martensi Karsch), a scorpion alpha-like toxin. The results indicate that hainantoxin-III and hainantoxin-IV are novel spider toxins and affect the mammal neural Na+ channels through a mechanism quite different from other spider toxins targeting the neural receptor site 3, such as delta-aractoxins and mu-agatoxins.
Xiao Y, et al. (2003) Inhibition of neuronal tetrodotoxin-sensitive Na+ channels by two spider toxins: hainantoxin-III and hainantoxin-IV. Eur J Pharmacol. PMID: 14512091
Structure and Function of Hainantoxin-III, a Selective Antagonist of Neuronal Tetrodotoxin-sensitive Voltage-gated Sodium Channels Isolated from the Chinese Bird Spider Ornithoctonus hainana
In the present study, we investigated the structure and function of hainantoxin-III (HNTX-III), a 33-residue polypeptide from the venom of the spider Ornithoctonus hainana. It is a selective antagonist of neuronal tetrodotoxin-sensitive voltage-gated sodium channels. HNTX-III suppressed Nav1.7 current amplitude without significantly altering the activation, inactivation, and repriming kinetics. Short extreme depolarizations partially activated the toxin-bound channel, indicating voltage-dependent inhibition of HNTX-III. HNTX-III increased the deactivation of the Nav1.7 current after extreme depolarizations. The HNTX-III·Nav1.7 complex was gradually dissociated upon prolonged strong depolarizations in a voltage-dependent manner, and the unbound toxin rebound to Nav1.7 after a long repolarization. Moreover, analysis of chimeric channels showed that the DIIS3-S4 linker was critical for HNTX-III binding to Nav1.7. These data are consistent with HNTX-III interacting with Nav1.7 site 4 and trapping the domain II voltage sensor in the closed state. The solution structure of HNTX-III was determined by two-dimensional NMR and shown to possess an inhibitor cystine knot motif. Structural analysis indicated that certain basic, hydrophobic, and aromatic residues mainly localized in the C terminus may constitute an amphiphilic surface potentially involved in HNTX-III binding to Nav1.7. Taken together, our results show that HNTX-III is distinct from β-scorpion toxins and other β-spider toxins in its mechanism of action and binding specificity and affinity. The present findings contribute to our understanding of the mechanism of toxin-sodium channel interaction and provide a useful tool for the investigation of the structure and function of sodium channel isoforms and for the development of analgesics.
Determination of disulfide bridges of two spider toxins: Hainantoxin-III and Hainantoxin-IV
Peptide toxins are usually highly bridged proteins with multipairs of intrachain disulfide bonds. Analysis of disulfide connectivity is an important facet of protein structure determination. In this paper, we successfully assigned the disulfide linkage of two novel peptide toxins, called HNTX-III and HNTX-IV, isolated from the venom of Ornithoctonus hainana spider. Both peptides are useful inhibitors of TTX-sensitive voltage-gated sodium channels and are composed of six cysteine residues that form three disulfide bonds, respectively. Firstly, the peptides were partially reduced by tris(2-carboxyethyl)-phosphine (TCEP) in 0.1 M citrate buffer containing 6 M guanidine-HCl at 40° C for ten minutes. Subsequently, the partially reduced intermediates containing free thiols were separated by reversed-phase high-performance liquid chromatography (RP-HPLC) and alkylated by rapid carboxamidomethylation. Then, the disulfide bonds of the intermediates were analyzed by Edman degradation. By using the strategy above, disulfide linkages of HNTX-III and HNTX-IV were determined as I-IV, II-V and III-VI pattern. In addition, this study also showed that this method may have a great potential for determining the disulfide bonds of spider peptide toxins.
Wang W, et al. (2009) Determination of disulfide bridges of two spider toxins: Hainantoxin-III and Hainantoxin-IV. J. Venom. Anim. Toxins incl. Trop. Dis