Liu Y, et al. (2012) A positively charged surface patch is important for hainantoxin-IV binding to voltage-gated sodium channels. J Pept Sci. PMID: 22927181

Li D, et al.(2004) Structure–activity relationships of hainantoxin-IV and structure determination of active and inactive sodium channel blockers. J Biol Chem. PMID: 15201273

A neurotoxin, named hainantoxin-IV, was purified from the venom of the spider Selenocosmia hainana. The amino acid sequence was determined by Edman degradation, revealing it to be a 35-residue polypeptide amidated at its C terminal and including three disulfide bridges: Cys2-Cys17, Cys9-Cys24, and Cys16-Cys31 assigned by partial reduction and sequence analysis. Hainantoxin-IV shares 80% sequence identity with huwentoxin-IV from the spider S. huwena, a potent antagonist that acts at site 1 on tetrodotoxin-sensitive (TTX-S) sodium channels, suggesting that hainantoxin-IV adopts an inhibitor cystine knot structural motif like huwentoin-IV. Under whole-cell voltage-clamp conditions, this toxin has no effect on tetrodotoxin-resistant voltage-gated sodium channels in adult rat dorsal root ganglion neurons, while it blocks TTX-S sodium channels in a manner similar to huwentoxin-IV, and the actions of both toxins on sodium currents are very similar to that of tetrodotoxin. Thus, they define a new family of spider toxins affecting sodium channels.

Liu Z, et al.(2003) Isolation and characterization of hainantoxin-IV, a novel antagonist of tetrodotoxin-sensitive sodium channels from the Chinese bird spider Selenocosmia hainana. Cell Mol Life Sci. PMID: 12827284

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

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.