Abstract The antimicrobial peptide ToAMP4 was isolated from Taraxacum officinale Wigg. flowers. Bacterial-expressed ToAMP4 showed antifungal activity but was inactive against bacteria. In this study, Chlamydomonas reinhardtii was used to express a C-terminal hemagglutinin (HA) and 6×His double tagged three repeats of ToAMP4 (3×ToAMP4)and the 3×ToAMP4-expressing cells were screened to express 3×ToAMP4 stably following continuous passaging for six months. Finally, a yield of 0.32 % of total soluble protein was achieved for 3×ToAMP4, which inhibits the growth of bacteria of both Gram-positive and Gram-negative at a minimum inhibitory concentration between 40 and 50 μg/ml. C. reinhardtii-derived 3×ToAMP4 maintained high stability in a wide range of temperature and pHs, tolerated to protease digestion at different extents and showed low hemolytic activity and cytotoxicity. Moreover, C. reinhardtii-expressed 3×ToAMP4 effectively caused damage on the membrane of targeted bacterial cells. In conclusion, our data show that C. reinhardtii has the potential to be used as a platform for the production of bioactive ToAMP4.

中文翻译：

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莱茵衣藻表达的 ToAMP4 多聚体抑制革兰氏阳性和革兰氏阴性细菌的生长

摘要 从蒲公英中分离得到抗菌肽ToAMP4。花卉。细菌表达的 ToAMP4 显示出抗真菌活性，但对细菌无活性。本研究采用莱茵衣藻表达C端血凝素（HA）和6×His双标记的ToAMP4（3×ToAMP4）3个重复序列，筛选出表达3×ToAMP4的细胞稳定表达3×ToAMP4。连续传代六个月。最后，3×ToAMP4 的总可溶性蛋白的产率为 0.32%，其以 40 至 50 μg/ml 的最小抑制浓度抑制革兰氏阳性菌和革兰氏阴性菌的生长。C. reinhardtii 衍生的 3×ToAMP4 在很宽的温度和 pH 范围内保持高稳定性，不同程度地耐受蛋白酶消化，并显示出低溶血活性和细胞毒性。此外，莱茵衣藻表达的 3×ToAMP4 有效地对目标细菌细胞的细胞膜造成损伤。总之，我们的数据表明，莱茵衣藻有可能用作生产生物活性 ToAMP4 的平台。