Jingzhaotoxin-III, a Novel Spider Toxin Inhibiting Activation of Voltage-gated Sodium Channel in Rat Cardiac Myocytes

Abstract

We have isolated a cardiotoxin, denoted jingzhaotoxin-III (JZTX-III), from the venom of the Chinese spider Chilobrachys jingzhao. The toxin contains 36 residues stabilized by three intracellular disulfide bridges (I-IV, II-V, and III-VI), assigned by a chemical strategy of partial reduction and sequence analysis. Cloned and sequenced using 3 -rapid amplification of cDNA ends and 5 -rapid amplification of cDNA ends, the full-length cDNA encoded a 63-residue precursor of JZTX-III. Different from other spider peptides, it contains an uncommon endoproteolytic site (-X-Ser-) anterior to mature protein and the intervening regions of 5 residues, which is the smallest in spider toxin cDNAs identified to date. Under whole cell recording, JZTX-III showed no effects on voltage-gated sodium channels (VGSCs) or calcium channels in dorsal root ganglion neurons, whereas it significantly inhibited tetrodotoxin-resistant VGSCs with an IC 50 value of 0.38 M in rat cardiac myocytes. Different from scorpion ␤-toxins, it caused a 10-mV depolarizing shift in the channel activation threshold. The binding site for JZTX-III on VGSCs is further suggested to be site 4 with a simple competitive assay, which at 10 M eliminated the slowing currents induced by Buthus martensi Karsch I (BMK-I, scorpion ␣-like toxin) completely. JZTX-III shows higher selectivity for VGSC isoforms than other spider toxins affecting VGSCs, and the toxin hopefully represents an important ligand for discriminating cardiac VGSC subtype.

Four Novel Tarantula Toxins as Selective Modulators of Voltage-Gated Sodium Channel Subtypes

“…Application of CcoTx1, CcoTx2, and PaurTx3 results in a shift of the current-voltage relationship of the studied VGSCs (Figs. 4 and 5 and Table 1), a characteristic that has also been reported for related spider toxins such as protoxin I, protoxin II, and Jingzhaotoxin III (Middleton et al, 2002;Xiao et al, 2004;Smith et al, 2005). In those studies, it was assumed that these toxins bind to the extracellular linker between DII-S3 and DII-S4 of the VGSC.…”

“…They reduce the current amplitude and have therefore been hypothesized to act by occluding the ion conduction pathway, similarly to toxins acting on site 1. Conversely, the protoxins and jingzhaotoxin-III shift the activation voltage toward more positive values, without affecting inactivation (Middleton et al, 2002;Xiao et al, 2004). The protoxins inhibit several VGSC subtypes but can also hamper the function of selected voltage-gated calcium or potassium channels.…”

Discovery of a selective Na V 1.7 inhibitor from centipede venom with analgesic efficacy exceeding morphine in rodent pain models

“…Rat DRG neurons were acutely dissociated and maintained in short-term primary culture, as previously described (31). Ca 2+ , Na + , and K + currents were recorded using the whole-cell patch-clamp technique with an Axon Multiclamp 700B amplifier (Molecular Devices).…”

SPider Neurotoxins Targeting Voltage-Gated Sodium Channels

“…with no change in inactivation kinetics (Xiao et al, 2004b). This depressant spider toxin showed no effects on Nav or Cav channels in DRG neurons or Kv channels expressed in Xenopus oocytes.…”