Purification and DNA binding properties of the ataxia-telangiectasia gene product ATM
Purification and DNA binding properties of the ataxia-telangiectasia gene product ATM
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DOI：
10.1073/pnas.96.20.11134
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发表时间：
1999-09-28
影响因子：
11.1
通讯作者：
Jackson, SP
中科院分区：
文献类型：
--
作者：
Smith, GCM;Cary, RB;Jackson, SP
人类神经退行性和癌症易感性疾病共济失调毛细血管扩张症在细胞水平上的特征是放射敏感性、染色体不稳定性和电离辐射诱导的细胞周期检查点控制受损。最近的研究表明，共济失调毛细血管扩张症的基因缺陷，被称为ATM，编码一个大约350 kda的多肽，ATM，是磷脂酰肌醇3-激酶家族的成员。我们证明了ATM结合DNA，并利用这一点来纯化ATM，使其接近均匀性。原子力显微镜显示，ATM存在于两个种群中，其大小与单体和四聚体状态一致。原子力显微镜分析也表明，atri优先结合DNA末端。这种特性类似于DNA依赖性蛋白激酶催化亚基所显示的特性，这是一个磷脂酰肌醇3-激酶家族成员，与DNA末端结合蛋白Ku一起在DNA损伤检测中起作用。此外，纯化的ATM含有一种激酶活性，以dna刺激的方式磷酸化p53的丝氨酸-15。这些结果为ATM提供了一个生化分析系统，支持了DNA依赖性蛋白激酶和ATM的不同作用的遗传数据，并提示了ATM如何指示p53和其他下游效应物DNA损伤的存在。
The human neurodegenerative and cancer predisposition condition ataxia-telangiectasia is characterized at the cellular level by radiosensitivity, chromosomal instability, and impaired induction of ionizing radiation-induced cell cycle checkpoint controls. Recent work has revealed that the gene defective in ataxia-telangiectasia, termed ATM, encodes an approximate to 350-kDa polypeptide, ATM, that is a member of the phosphatidylinositol 3-kinase family. We show that ATM binds DNA and exploit this to purify ATM to near homogeneity. Atomic force microscopy reveals that ATM exists in two populations, with sizes consistent with monomeric and tetrameric states. Atomic force microscopy analyses also show that ATR I binds preferentially to DNA ends. This property is similar to that displayed by the DNA-dependent protein kinase catalytic subunit, a phosphatidylinositol 3-kinase family member that functions in DNA damage detection in conjunction with the DNA end binding protein Ku. Furthermore, purified ATM contains a kinase activity that phosphorylates serine-15 of p53 in a DNA-stimulated manner. These results provide a biochemical assay system for ATM, support genetic data indicating distinct roles for DNA-dependent protein kinase and ATM, and suggest how ATM may signal the presence of DNA damage to p53 and other downstream effectors.