黃麴毒素調控蛋白質AflR之免疫化學及分子功能性研究(II)

中山醫學大學機構典藏 CSMUIR > 醫學科技學院 > 生物醫學科學學系暨碩士班 > 研究計劃 > Item 310902500/2703

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題名:	黃麴毒素調控蛋白質AflR之免疫化學及分子功能性研究(II) Immunochemical and Molecular Studies of the Regulatory Protein, AflR, Involved in Aflatoxin Biosynthesis (II)
作者:	劉秉慧 Liu, Biing-Hui
貢獻者:	中山醫學院生命科學系
關鍵詞:	黃麴毒素;調節性蛋白質AFLR;生合成;免疫化學;去氧核糖核酸結合能力 Aflatoxin；Regulatory protein AFLR；Biosynthesis；Immunochemistry；DNA binding ability
日期:	2002
上傳時間:	2010-11-05T10:48:50Z (UTC)
摘要:	黃麴毒素是一族致癌及致突變的真菌毒素，為了瞭解在黃麴毒素的生化合成途徑中，調控基因AflR所扮演的角色，首先將aflR的cDNA在大腸桿菌中表達成為AflR蛋白質，並且以此為Balb/c老鼠的免疫原產生五株穩定的融合瘤細胞株並且予以定性。此外，利用特定序列的DNA親和層析法將分子量47.5 kDa的AflR蛋白質自產毒A. parasiticus菌株中純化出來；純化後的蛋白質能夠與兩段分別位於aflR基因和黃麴毒素合成基因的啟動區域在活體外形成複合物；然而來自於非產毒菌株的A. oryzae的蛋白質萃取物則完全無法辨識該兩段區域；研究結果顯示在黃麴菌屬中。AflR與特定DNA區域的結合能力與菌株是否會產生黃麴毒素的特性應有直接的相關性。 To investigate the role of regulatory protein AflR in aflatoxin biosynthetic pathway, recombinant AflR was expressed in E.coli and purified homogeneously. With recombinant AflR as immunogen, five stable hybrodoma cell lines were generated and characterized. On the other hand, the native AflR was purified from A. parasiticus by sequence-specific DNA affinity chromatography. Purified AflR with a molecular weight of 47.5 kDa not only bound the AflR binding site (5'-TTAGGCCTAA-3') found upstream of the aflR gene, but also bound the sequence II (5'-TCGNNNNNCGA-3') in the promoter region of the structural genes. In contrast, partially purified protein extracts from nonaflatoxigenic A. oryzae did not recognize either of the DNA sequences as compared to aflatoxigenic A. parasiticus AflR in electrophoretic mobility shift assays. These findings suggest that the sequence-specific DNA binding ability of AflR is essential for the activation of aflatoxin biosynthetic pathway.
URI:	https://ir.csmu.edu.tw:8080/handle/310902500/2703
顯示於類別:	[生物醫學科學學系暨碩士班] 研究計劃

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