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Title: 抗氧化相關脢cDNA之選殖、表現、特性及其動力學之研究
Cloning, Expression, Properties, and Kinetic Studies of Antioxidant Enzymes
Authors: 林棋財
Contributors: NTOU:Institute of Bioscience and Biotechnology
Keywords: 樟芝子實體;抗氧化相關?;酵母菌
Antrodia camphorata;antioxidant enzyme;yeast
Date: 2008-08
Issue Date: 2011-06-28T07:17:49Z
Publisher: 行政院國家科學委員會
Abstract: 摘要:抗氧化相關酶包含GFD (glutathione-dependent formaldehyde dehydrogenase), GPx (glutathione peroxidase), DHAR (dehydroascorbate reductase), MDAR (monodehydroascorbate reductase), TR (thioredoxin reductase), Grx (glutaredoxin), SOD (superoxide dismutase), Prx (peroxiredoxin) 包含1-Cys Prx 和2-Cys Prx, Trx (thioredoxin), ascorbate peroxidase, metallothionin 等,對維持身體健康扮演重要角色。以新鮮樟芝子實體為材 料,抽取RNA,合成cDNA,建立更大EST,藉由EST 序列資訊,用樟芝子實 體cDNA 為模板,以PCR 增生抗氧化相關酶DNA 並進行篩選,應可篩得全長 抗氧化相關酶之cDNA,繼之與表現型載体連接,送入E.coli 和酵母菌進 行大量生產並純化。純化之相關酶進行生化性質測試及其動力學之研究。 抗氧化相關酶均為蛋白質,如何進入真核細胞是一大考驗,因此擬在N 端 分別接上幾個胺酸(Tat),則Tat-抗氧化相關酶產物將容易進入細胞以發揮 其功能。純化得到之Tat-抗氧化相關酶產物將進行應用實驗。所得如為正 面效果,將申請專利並與生技公司合作應用於化妝品,健康食品或燙傷藥 膏中。
Abstract:Antioxidant enzymes include GFD (glutathione-dependent formaldehyde dehydrogenase), GPx (glutathione peroxidase), DHAR (dehydroascorbate reductase), MDAR (monodehydroascorbate reductase), TR (thioredoxin reductase), Grx (glutaredoxin), SOD (superoxide dismutase), Prx (peroxiredoxin)including 1-Cys Prx and 2-Cys Prx, Trx (thioredoxin), ascorbate peroxidase. These enzymes are useful for living cell. Based on cDNA library and EST from the fresh fruit body of Antrodia camphorata , hoping more antioxidant enzyme cDNAs will be cloned, subsequently the active enzymes will be obtained from bacteria or yeast.In addition, we will study its biochemical and kinetic properties. Possibly these antioxidant enzymes may be used for burn healing agents or cosmetics for the protection of the skin, or unesthetic effects caused by peroxides, free radicals. Antrodia camphorata have physiological activities, especially the stimulation of the immune system and the prevention effects on cancer. They are reported having the capabilities to stimulate the proliferation of immune cells, increase the production of cytokines and tumor necrosis factor-γ. Antrodia camphorata, one kind of the medicinal mushrooms, can naturally grow in the hay of Cinnamomum kanehirai that is a special camphor tree locally existing in Taiwan. This encourages us to clone full-length cDNA of antioxidant enzymes from A. camphorata, then are introduced into expression vectors, we hope to obtain large amount of high activity and stable enzymes. The molecular mass of antioxidant enzymes were estimated to be 41-16 kDa, is not easily transferred into mammalian cell, if fused with Tat on N-terminus of those proteins, the barrier will be solved.
Relation: NSC97-2313-B019-001-MY3
URI: http://ntour.ntou.edu.tw/ir/handle/987654321/9885
Appears in Collections:[生命科學暨生物科技學系] 研究計畫

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