Cloning of Drosophila Gene which Code for UV-damaged DNA Binding Protein.

编码紫外线损伤 DNA 结合蛋白的果蝇基因的克隆。

• 批准号: 01580211
• 负责人: TODO Takeshi
• 金额: $ 1.34万
• 依托单位: Department of Fundamental Biology, Osaka University Medical School
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1989
• 资助国家: 日本
• 起止时间: 1989 至 1990
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-01580211/
• 关键词: DNA repair; Ultrayiolet light; Gel Shift Assay; Drosophila; DNA binding protein; 紫外線損傷; ゲルシフト法

Using a gel electrophoresis DNA band shift assay, we have identified two DNA binding protein complex in Drosophila embryonic cell which have a high affinity for UV-irradiated, double-stranded DNA. Screening Drosophila mutants which are deficient in DNA repair has identified five mutants which lack either of the two protein complex. Four excision repair deficient mutants (mus201, phr, mus308 and mus205) lack one protein complex (Factor 2). Another protein complex (Factor 1) was not detectable in post-replication repair deficient mutant mus104. These findings suggest a possible involvement of t he gene product in lesion recoghition and DNA repair of UV-light induced photoproduct.Using heparin agarose column chromatography, we can separate each factor into different fraction. The fraction which contain Factor 2, that is the fraction which have ability to form band 2, was further purified by UV-irradiated DNA affinity column chromatography. In the fraction which has ability to form band 2, we can identify only one protein band in silver stained SDS-PAGE gel. The molecular mass was assummed to be between 45k and 50k dalton. We assumed this protein is Factor 2.

用凝胶电泳DNA带移分析法，我们在果蝇胚胎细胞中鉴定出两个DNA结合蛋白复合物，它们对紫外线照射的双链DNA具有高亲和力。通过筛选果蝇DNA修复缺陷突变体，鉴定出5个缺失这两种蛋白复合物中任一种的突变体。四个切除修复缺陷突变体（mus 201，phr，mus 308和mus 205）缺乏一个蛋白质复合物（因子2）。另一种蛋白复合物（因子1）在复制后修复缺陷突变体mus 104中检测不到。这些结果表明，该基因产物可能参与了紫外光诱导的光产物的损伤识别和DNA修复。含有因子2的级分，即具有形成条带2的能力的级分，通过UV照射的DNA亲和柱层析进一步纯化。在能够形成条带2的组分中，我们在银染SDS-PAGE凝胶上只能鉴定出一条蛋白条带。分子量在45 k和50 k道尔顿之间。我们假设这种蛋白质是因子2。

项目成果

Takeshi Todo and Haruko Ryo: "Identification of the Protein Factors which Bind Specifically to Ultraviolet Light Irradiated DNA." J. Radiat. Res. 31. 26 (1990)

Takeshi Todo 和 Haruko Ryo：“鉴定与紫外线照射的 DNA 特异性结合的蛋白质因子。”

Takeshi Toda and Haruko Ryo: "Identification of cellular factors that recognize DNA containing UV damage in Drosophila melanogaster." Mutation Research.

Takeshi Toda 和 Haruko Ryo：“识别黑腹果蝇中含有紫外线损伤 DNA 的细胞因子的鉴定。”

Takeshi Todo: "Detection of cellular factors that bind to UVーdamaged DNA in Drosophila" Mutation Research.

Takeshi Todo：“检测果蝇中与紫外线损伤 DNA 结合的细胞因子”突变研究。

Takeshi Todo and Haruko Ryo: "Identification of the protein factors which bind specifically to Ultraviolet light irradiated DNA." Journal of Radiation Resarch. 31. 26 (1990)

Takeshi Todo 和 Haruko Ryo：“鉴定与紫外线照射的 DNA 特异性结合的蛋白质因子。”

Takeshi Todo and Haruko Ryo: "Identification of cellular factors that recognize DNA containing UV damage in Drosophila melanogaster." Mutation Research.

Takeshi Todo 和 Haruko Ryo：“识别黑腹果蝇中含有紫外线损伤 DNA 的细胞因子的鉴定。”