[Synthesis and oxidative refolding of an N-terminal truncate of jingzhaotoxin-V and characterization of its activities of sodium channels].
An N-terminal tyrosine residue truncate of Jingzhaotoxin-V (Y1-JZTX-V) was synthesized by solid-phase chemical methods using Fmoc-protected amino acids. Reversed-phase high performance liquid chromatography (RP-HPLC) and matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF/TOF MS) were used to monitor the oxidative refolding of Y1-JZTX-V to find the optimal renaturation conditions of the synthetic linear peptide. When Y1-JZTX-V (0.05 mg/L) was dissolved in 0.1 mol/L Tris-HCl buffer containing 1 mmol/L GSH and 0.1 mmol/L GSSG at pH 7.50 and 4 degrees C, the best renaturation yield of the truncate toxin was obtained. Under the whole-cell patch-clamp mode, Y1-JZTX-V could inhibit tetrodotoxin-resistant (TTX-R) and tetrodotoxin-sensitive (TTX-S) sodium currents in adult rat dorsal root ganglion neurons with IC50 values of 160 nmol/L and 39.6 nmol/L, respectively. The inhibition potentiality of Y1-JZTX-V on TTX-S sodium currents was almost the same as the natural JZTX-V, while that on TTX-R sodium currents was obviously weakened. The IC50 value of Y1-JZTX-V on TTX-R sodium currents was 5.8 times as many as that of natural JZTX-V. Present findings indicated that the first tyrosine residue (Y1) in the N-terminal of JZTX-V was involved in the binding activities of JZTX-V to TTX-R sodium channels.
Quan
Miaohua
M
The Key Laboratory of Protein Chemistry and Developmental Biology of Ministry of Education, Hunan Normal University, Changsha 410081, China.
Zeng
Xiongzhi
X
Pi
Jianhui
J
Deng
Meichun
M
Liang
Songping
S
D002851:Chromatography, High Pressure Liquid; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology*; D017510:Protein Folding; D015222:Sodium Channels / Q000187:drug effects*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013111:Spider Venoms / Q000737:chemistry*
D010455:Peptides; D015222:Sodium Channels; D013111:Spider Venoms; C518577:jingzhaotoxin-V, Chilobrachys jingzhao