Determination of unknown poly(ADP-ribosyl)ated proteins and binding sites

未知聚（ADP-核糖基）化蛋白和结合位点的测定

• 批准号: 16390072
• 负责人: MIWA Masanao
• 金额: $ 7.94万
• 依托单位: Nagahama Institute of Bio-Science and Technology (2005)University of Tsukuba (2004)
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2004
• 资助国家: 日本
• 起止时间: 2004 至 2005
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-16390072/
• 关键词: Posttranslational modification; Poly(ADP-ribosyl)ation; Poly(ADP-ribose) glycohydrolase; Drosophila melanogaster; PARP; Centrosome; ポリADP-リボース; p53; ポリADP-リボース分解酵素; 神経変性; 細胞周期; 細胞分裂

We identified tow proteins that are poly(ADP-ribosyl)ated in vivo. They are p53 and nucleophosmin/B23. To identify the site of poly(ADP-ribosyl)ation of p53, we made the expression vector to produce GST-p53 fusion protein in E. coli. We confirmed the poly(ADP-ribosyl)ation of the entire product of p53 (aa1-aa393) by PARP-1. Currently we are preparing several GST-truncated p53 proteins, that lack about 40 amino acids, and analyzing poly(ADP-ribosyl)ated domain of p53.We identified a GST-truncated p53 protein, in which about 40 amino acids are deleted in the DNA binding domain, that lacks the acceptor activity of poly(ADP-ribosyl)ation. To identify the poly(ADP-ribosyl)ated proteins in general, we made an affinity column that have the antibody against poly(ADP-ribose), 1OH. We are optimizing the condition to bind the proteins from centrosome fractions and from other cellular fractions. To prepare a large amount of proteins that are poly(ADP-ribosylated), we are collecting the proteins from Drosophila mutant that lacks poly(ADP-ribose) glycohydrolase.

我们在体内鉴定了两种聚腺苷二磷酸核糖结合的蛋白质。它们是P53和核磷蛋白/B23。为了确定P53多聚(ADP-核糖基)的位置，我们构建了表达载体，在大肠杆菌中表达了GST-P53融合蛋白。我们通过PARP-1证实了P53(Aa1-aa393)的整个产物的聚(ADP-核糖基)。目前，我们正在制备几种缺失约40个氨基酸的GST缺失的p53蛋白，并对p53的聚(ADP-核糖基)结构域进行分析。我们发现了一个缺失约40个氨基酸的缺失DNA结合区的GST-缺失的p53蛋白，它缺乏聚(ADP-核糖基)的受体活性。为了从总体上鉴定聚腺苷二磷酸核糖结合蛋白，我们制作了一种亲和层析柱，其抗体为聚腺苷二磷酸核糖1OH。我们正在优化条件，以结合来自中心体部分和其他细胞部分的蛋白质。为了制备大量聚腺苷二磷酸核糖化的蛋白质，我们收集了缺乏聚腺苷二磷酸核糖水解酶的果蝇突变体的蛋白质。

项目成果

Integration of human T-cell leukemia virus type 1 in genes of leukemia cells of patients with adult T-cell leukemia

• DOI: 10.1111/j.1349-7006.2004.tb03207.x
• 发表时间: 2004-04-01
• 期刊: CANCER SCIENCE
• 影响因子: 5.7
• 作者: Hanai, S;Nitta, T;Miwa, M
• 通讯作者: Miwa, M

New development of research on poly (ADP-ribosyl) ation (in Japanese)

聚（ADP-核糖基）化研究新进展（日文）

• 发表时间: 2004
• 期刊: Jikken Igaku 22(9)
• 作者: Miwa;M.;Kanai;M.;Hanai;S.
• 通讯作者: S.

Loss of poly(ADP-ribose) glycohydrolase causes progressive neurodegeneration in Drosophila melanogaster

• DOI: 10.1073/pnas.2237114100
• 发表时间: 2004-01-06
• 期刊: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
• 影响因子: 11.1
• 作者: Hanai, S;Kanai, M;Miwa, M
• 通讯作者: Miwa, M

ポリ(ADP-リボース)代謝と細胞機能

聚（ADP-核糖）代谢和细胞功能

• 发表时间: 2006
• 期刊: 生化学(日本生化学会誌 : 総説) 78(In press)
• 作者: 三輪 正直;益谷 美都子
• 通讯作者: 益谷 美都子

Cellular function by poly(ADP-ribosyl)ation, especially the regulation of centrosome function and neuronal cells (in Japanese)

聚（ADP-核糖基）化的细胞功能，特别是中心体功能和神经元细胞的调节（日语）

• 发表时间: 2004
• 期刊: Vitamin 78(7)
• 作者: Miwa;M.;Kanai;M.;Sugihara;E.;Ohashi;S.;Nogi;Y.;Hanai;S.
• 通讯作者: S.