天麻免疫調節蛋白之純化與活性分析

Purification and Function Analysis of the Immunomodulatory Protein from Gastrodia elata

陳美如(Mei-Ju Chen)

指導教授 ： 許輔

國立臺灣大學/生物資源暨農學院/園藝暨景觀學系/碩士(2015年)

https://doi.org/10.6342/NTU.2015.10442

摘要

無資料

關鍵字

天麻 ； 純化 ； 免疫調節蛋白

並列摘要

Tianma (Gastrodia elata), also known as fixed wind grass or barbed arrows, is a medicinal orchid used in traditional Chinese medicine to treat vertigo, paralysis, convulsions, cramps, epilepsy, high blood pressure and other symptoms. We have previously purified an immunomodulatory protein IPAF from Anoectochilus formosanus. The protein IPAF has an amino acid sequence highly similar to proteins from Orchidaceae plants, including Epipactis helleborine lectin, Listera ovata lectin, and G. elata-gastrodianin-4B. Therefore, we hypothesized that Orchidaceae plants might possess proteins similar to IPAF. To test our hypothesis, the crude protein was extracted from G. elata, Dendrobium huoshanense, Dendrobium nobile, and Dendrobium loddigesii and then analyzed by polyacrylamide gel electrophoresis and Western blotting. Subsequently, a 15.1 kDa protein G. elata (GEP), which exhibited similar biochemical characteristics to IPAF, was purified via anion exchange chromatography using DE-52 and HiTrap Q columns. Finally, RAW 264.7 murine macrophage cells were used for preliminary activity tests. GEP (5 μg/mL) could activate nitric oxide (NO) (10.4 ± 0.6 μM), TNF-α (935.4 ± 118.5 pg/mL), and IL-1β (25.1 ± 3.0 pg/mL) secretion by RAW 264.7 murine macrophage. In addition, GEP enhanced nucleic acid replication by 1.16-fold as analyzed by BrdU analysis, and promoted 1.43-fold of MTT metabolic activity in murine splenocytes. In this study we purified a new orchid protein, which exhibited similar biochemical characteristics as the IPAF, and we also assessed its immunomodulatory function. GEP activated RAW 264.7 macrophages by enhancing NO, TNF-α and IL-1β secretion. Moreover, GEP also stimulated cell proliferation in mouse splenocytes. On the basis of previous the findings, in our laboratory GEP, as well as other orchid proteins, might play important roles in the medicinal functions of Orchidaceae plants.

並列關鍵字

Gastrodia elata ； Purification ； Immunomodulatory Protein

參考文獻

李婉慈（民99）。探討台灣金線連免疫調節蛋白對小鼠腹腔巨噬細胞之活化作用

林世航（民101）。以蛋白質體學探討天麻水萃物抗憂鬱之功效（未出版之碩士論

許如君（民94）。台灣金線連免疫調節蛋白 IPAF 活化免疫細胞之機制（未出版

Ahn, E. K., Jeon, H. J., Lim, E. J., Jung, H. J., Park, E. H. (2007). Anti- inflammatory and anti-angiogenic activities of Gastrodia elata Blume. J Ethnopharmacol, 110, 476-482.

An, S. J., Park, S. K., Hwang, I. K., Choi, S. Y., Kim, S. K., Kwon, O. S., Jung, S. J., Baek, N. I., Lee, H. Y., Won, M. H., Kang, T. C. (2003). Gastrodin decreases