Studies On The Structure And Function Of Tow Spider Toxins

Jingzhaotoxin-Ⅴ(chilobrachys jingzhao,JZTX-Ⅴ) is an inhibitor cystine knot toxin that selectively inhibit wild Kv4.3 expressed in HEK293 and the IC50 value is 10nmol/L.Electrophysiological assays indicated that 25nmol/L JZTX-Ⅴcould shift significantly the voltage dependence of activation and steady-state inactivation to depolarization,and slow markedly the time constant of activation and inactivation but not the recovery from inactivation.Thus,JZTX-Ⅴis likely a gating modifier toxin.To study the molecular determinants of interaction,we performed alanine scanning on the S3b—S4 region in Kv4.3 voltage-sensing domain and researched the interaction between mutants and JZTX-Ⅴ.The results showed that V282 and F286 were molecular determinants of the JZTX-Ⅴon the Voltage-sensing Domains of Kv4.3.Besides,in order to observe the effects of Jingzhaotoxin-V(JZTX-V) on withdrawl symptoms which was aroused by Naloxone precipitation,the animal model of morphine dependent in mice was established.The results indicated that during the 15min after withdrawl,0.1μg/20g,1μg/20g and 10μg/20g JZTX-Ⅴcould significantly reduce jumping times(P<0.05 or 0.01).According to these phenomenon and data,JZTX-Ⅴcould become a leading molecule that could be used for getting rid of drugs.Using cation exchange chromatography and reverse phase high performce liquid chromatography,a novel neurotoxin named Raventoxin-Ⅶwas purified from the crude venom of Macrothele raveni.The molecular weight of Raventoxin-Ⅶis 3158.155Da,identified by MALDI-TOF mass spectrometry.Raventoxin-Ⅶcontains three disulfide bonds which are made up of six cysteines from its complete amino acid seqence determined by automatic Edman degradation.The preliminary electrophysiological results showed that Raventoxin-Ⅶcould inhibit voltage-gated TTX-sensitive sodium channel current(IC50=12.2μmol/L),but not the voltage-gated TTX-resistant sodium channel current.