Recombinant Expression, Functional Characterization of Two Scorpion Venom Toxins with Three Disulfide Bridges from the Chinese Scorpion Buthus martensii Karsch

Abstract

Scorpion venom contains a large variety of biologically active peptides. However, most of these peptides have not been identified and characterized. Peptides with three disulfide bridges, existing in the scorpion venom, have not been studied in detail and have been poorly characterized until now. Here, we report the recombinant expression and functional characterization of two kinds of venom peptides (BmKBTx and BmNaL-3SS2) with three disulfide bridges. This study adopted an effective Escherichia coli system. The genes for BmKBTx and BmNaL-3SS2 were obtained by polymerase chain reaction and cloned to the pSYPU-1b vector. After expression and purification, the two recombinant proteins were subjected to an analgesic activity assay in mice and whole-cell patchclamp recording of hNav1.7-CHO cell lines. Functional tests showed that BmKBTx and BmNaL- 3SS2 have analgesic activity in mice and can interact with the hNav1.7 subtype of the voltage-gated sodium channel (VGSC). Scorpion venom is rich in bioactive proteins, but most of their functions are unknown to us. This study has increased our knowledge of these novel disulfide-bridged peptides (DBPs) and their biological activities.

Peptide Inhibitors of Kv1.5: An Option for the Treatment of Atrial Fibrillation

“…One of the main reasons for the lack of peptide inhibitors for Kv1.5 is the presence of a positively charged Arg residue (R487) in the pore region of the channel, a feature missing in other Kv channels [103,104], which prevents Kv channel pore blocker peptides from binding to Kv1.5. The mutation of this Arg to Val (R487V) or Tyr (R487Y) made Kv1.5 sensitive to BgK, a known inhibitor toxin for Kv1 channels from the sea anemone Bunodosoma granulifera [103,105,106]. Knowing the binding mechanism of other toxins to Kv channels and the particular properties of the Kv1.5 channel, the selectivity of Ts6 and Osu1 (and other peptides) and their affinity for Kv1.5 could be improved by rational drug design in which their amino acid sequences are modified in a targeted way guided by in silico docking experiments.…”

“…This problem can be solved by chemical synthesis or recombinant expression of these peptides. After appropriate standardization of the expression protocols, a sufficient amount of peptide can be obtained [105][106][107] to fully characterize the biological activity of the peptides. However, as mentioned earlier, most toxin peptides are rich in disulfide bonds, resulting in low yields of the bioactive product with the desired disulfide bridge configuration.…”

Scorpion inhibits epithelial–mesenchymal transition and metastasis of hepatocellular carcinoma

“…There are many active ingredients in Scorpion, including antimicrobial peptides Mucroporin-M1, 31 analgesic peptide BmKBTx, 32 antiepilepsy peptide AEP, 33 antitumor peptide HS-1, 34 etc. We only detected antitumor effect of Scorpion; however, it remains unclear which active ingredient has the role in inhibiting tumor EMT and metastasis.…”

Cloning, expression and identification of KTX-Sp4, a selective Kv1.3 peptidic blocker from Scorpiops pococki

“…Since there are fewer kinds of scorpion species being used to discover bioactive peptides, only about 0.4–0.5% of toxin peptides have been identified and clearly characterized so far [ 24 ]. Till now, the reported toxin polypeptide extracted from Xizang Scorpiops pococki has been far less than that from other scorpions such as Buthus martensii Karsch [ 25 , 26 ]. By combined use of transcriptome sequence and functional gene prediction in present work, the KTX - Sp4 gene in the cDNA Library of Xizang Scorpiops pococki was screened, and then successfully expressed in prokaryotic expression system.…”