Oxidatively damaged DNA Repair and Its Related Enzymes

氧化损伤DNA修复及其相关酶

• 批准号: 13480193
• 负责人: KURAMITSU Seiki
• 金额: $ 9.28万
• 依托单位: Osaka University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-13480193/
• 关键词: DNA repair; damaged DNA; nucleotide; catalytic mechanism; substrate recognition; extreme thermophile; 酸化傷害DNA修復; 8-オキソグアニン; MutM; フリップアウト; Thermus thermophilus HB8; 立体構造反応解析; 基質特異性; 分子機能解析

DNA is oxidatively damaged by oxidative stress especially reactive oxygen species. Such damages result in mutations and cause a number of diseases including cancer. Among oxidatively damaged bases 8-oxoguanine can lead to increased frequency of G : C to T : A transversion. MutM is a base excision repair enzyme that recognize oxidatively damaged guanines including 8-oxoguanine. This enzyme catalyzes three reactions: excision of the 8-oxoguanine base (glycosylase activity). and b-and d-elimination of the resultant abasic site (AP-lyase activity), Recently, we also determined the structures of MutM-DNA complex. These structures suggest involvement of many amino acid residues in DNA-binding and catalytic activities. However, proposed roles of those residues have not been verified experimentally. To establish the detailed mechanism of MutM reaction, this study investigated functional roles of several amino acid residues in the catalysis. We selected several conserved residues based on the crystal structure of the complex and replaced them by other residues using site-directed mutagenesis. The obtained results demonstrate that some residues are involved in the catalytic reaction of MutM.

氧化应激尤其是活性氧对DNA有氧化损伤作用。这种损害会导致突变并导致包括癌症在内的多种疾病。在氧化损伤的碱基中，8-氧代鸟嘌呤可导致G：C到T：A颠换频率增加。MutM是一种碱基切除修复酶，识别氧化损伤的鸟嘌呤，包括8-氧代鸟嘌呤。该酶催化三个反应：切除8-氧代鸟嘌呤碱基（糖基化酶活性）。以及由此产生的脱碱基位点的b-和d-消除（AP-裂解酶活性）。最近，我们还测定了MutM-DNA复合物的结构。这些结构表明许多氨基酸残基参与DNA结合和催化活性。然而，这些残留物的拟议作用尚未得到实验验证。为了建立MutM反应的详细机理，本研究考察了几个氨基酸残基在催化中的功能作用。我们根据复合物的晶体结构选择了几个保守的残基，并使用定点突变将其替换为其他残基。所得结果表明，一些残基参与MutM的催化反应。

项目成果

Masui, R.: "MutM : Single C_2C_2 Zinc Finger-DNA interaction"Landes Bioscience(印刷中).

Masui, R.：“MutM：单 C_2C_2 锌指-DNA 相互作用”Landes Bioscience（正在出版）。

Masui, R.: "Zinc Finger Proteins : from Atomic Contact to Cellular Function (edited by Iuchi, S. and Kuldell.N.)"Landes Bioscience(印刷中). (2004)

Masui, R.：“锌指蛋白：从原子接触到细胞功能（由 Iuchi, S. 和 Kuldell.N. 编辑）”Landes Bioscience（2004 年出版）。

Yamagata, A.: "Overexpression, Purification, and Characterization of Rec.J Protein from Thermus thermophilus HB8 and Its Core Domain"Nucleic Acids Res.. 29(22). 4617-4624 (2001)

Yamagata, A.：“来自嗜热栖热菌 HB8 的 Rec.J 蛋白及其核心结构域的过表达、纯化和表征”核酸研究 29(22)。

Masui, R., Nakagawa.N., Kuramitsu, S.: "MutM: Single C_2C_2 Zinc Finger-DNA interaction"Zinc Finger Proteins: from Atomic Contact to Cellular Function (edited by Iuchi, S. Kuldell, N.) Landes Bioscience. (in press).

Masui, R.、Nakakawa.N.、Kuramitsu, S.：“MutM：单 C_2C_2 锌指-DNA 相互作用”锌指蛋白：从原子接触到细胞功能（由 Iuchi, S. Kuldell, N. 编辑）Landes Bioscience

Iwai, T.: "The Nudix Hydrolase Ndx1 from Thermus thermophilus HB8 Is a Diadenosine Hexaphosphate Hydrolase with a Novel Activity"J.Biol.Chem.. in press. (2004)

Iwai, T.：“来自嗜热栖热菌 HB8 的 Nudix 水解酶 Ndx1 是一种具有新颖活性的二腺苷六磷酸水解酶”J.Biol.Chem.. 正在出版。