台灣產虎頭蜂蜂毒胜肽(mastoparans)之生物特性

Mastoparan is the most abundant peptide in the hornet venoms. The aim of this study is to investigate biological characteristics of mastoparans isolated from Vespa species in Taiwan, i.e., Vespa affinis, Vespa analis, Vespa basalis, Vespa ducalis, Vespa mandarinia and Vespa velutina. Using PCR to amplify mastoparans cDNA fragments with specific primers, six cDNA sequences encoding mastoparan precursors were cloned and named mastoparan-AF, mastoparan-A, mastoparan-B, mastoparan-D, mastoparan-M and mastoparan-V, respectively. Among six mastoparans, mastoparan-D and mastoparan-V are novel genes. After translation, the precursors of these mastoparans are composed of N-terminal signal sequence, prosequence, mature peptide and appendix C-terminal glycine residue. Subsequently, six mastoparans were synthesized for studying their biological characteristics. The circular dichroism spectra of mastoparans showed unordered random coil in water and a high content α-helical conformation in the presence of 8 mM sodium dodecyl sulfate (SDS) and 40% 2,2,2-trifluoroethanol (TFE). In mast cell degranulation assay, mastoparans caused mast cell degranulation in Sprague-Dawley (SD) rats. In antimicrobial activity assay, mastoparans exhibited antimicrobial activity against both Gram-positive and Gram-negative bacteria. Mastoparan-AF exhibited more potent antimicrobial activity than others. In membrane permeabilization, mastoparans caused dramatically membrane permeabilization on Escherichia coli BL21 with the increase of mastoparan concentrations. In hemolytic activity assay, mastoparans caused slight hemolysis on human and chicken erythrocytes but almost no hemolysis on goat erythrocytes at effective antimicrobial concentrations. Our results also showed that mastoparan-AF exhibited more potent antimicrobial activity than clinically used antibiotics against E. coli isolates. Furthermore, mastoparan-AF in combination with certain antibiotics, i.e., cephalothin or gentamicin, resulted in synergistic antimicrobial activity against multiple-antibiotic-resistant E. coli isolates, i.e., E. coli PFH13. As mentioned above, mastoparan-AF alone or in combination with antibiotic could exhibit antimicrobial activity. It is also revealed that mastoparan-AF could be an alternative for conventional antibiotics and is worth further developing as an antimicrobial medicine.