Abstract:

Jingzhaotoxin-I (JZTX-I), a 33-residue polypeptide with three disulfide bonds, was a novel spider neurotoxin preferentially inhibiting cardiac sodium channel inactivation. Its activities of phospholipid membrane-binding were studied by a combination of reversed-phase high performance liquid chromatography (HPLC) and fluorescence spectroscopy. Small unilamellar vesicles binding assays showed that the partitioning of JZTX-I into lipid bilayer did not require negatively charged phospholipids. Further, JZTX-I also exhibited a blue shift of 6.4 nm or 4.7 nm as well as red-edge excitation shift of 7.4 nm or 8.0 nm in the presence of 75% 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE)/25% 1-palmitoyl-2-oleoyl-sn-glycero-3-［phospho-rac-(1-glycerol)］ (sodium salt) (POPG) or 100% 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) vesicles respectively, suggesting that some tryptophan residues on the hydrophobic surface of the toxin were located within a motion restricted membrane interfacial region. Fluorescence quenching experiments suggested that some tryptophan residues of JZTX-I were positioned within the membrane and protected from aqueous quenching agents. These findings should provide further insight into the molecular mechanism of the channel gating of JZTX-I.

Key words: fluorescence spectroscopy, Jingzhaotoxin-I (JZTX-I) , small unilamellar vesicles, high performance liquid chromatography (HPLC)