FORMATION, MUTAGENICITY & REPAIR OF PHOTOHYDRATES IN DNA

我们将确定碱基切除修复氧化损伤的DNA 碱基优先出现在活跃转录的基因中。修理费 5-羟甲基尿嘧啶(HmUra)，由HmUra-DNA糖基酶修复 且不阻断复制，将其与胸腺嘧啶二醇进行比较 以及由大肠杆菌核酸内切酶III和DO修复的相关损伤 数据块复制。HmUra-DNA糖基酶在修复酶中是独一无二的 没有细菌和酵母的。我们已经提议它的 系统发育分布反映了它的主要功能，即修复 DNA中的5-甲基胞嘧啶残留量。相反，类核酸内切酶III 活动在系统发育上是保守的，这表明损伤 被酶修复的是在所有物种中形成的。在这些损伤中有 紫外线诱导的水合嘧啶。胞嘧啶的水合作用导致 尿嘧啶水合物的不可逆脱氨反应，如果不修复，会导致 点突变。该反应方案也可用于5-甲基胞嘧啶 导致不可逆脱氨生成水合胸腺嘧啶的残留物。我们会 水溶液中嘧啶类光水合物的形成及稳定性研究 利用大肠杆菌内切酶制备高比活度寡核苷酸 III作为探头。我们将使用这些寡核苷酸作为底物 哺乳动物核酸内切酶Ⅲ类活性的纯化 足以获得蛋白质序列和分子克隆。结果是 这些实验将增加我们对形成和修复的知识 对DNA的氧化和光化学损伤，并可能阐明 这种损害与光化性和自发性的病因学关系 癌症。

We will determine whether base excision repair of oxidatively damaged DNA bases occurs preferentially in actively transcribed genes. The repair of 5-hydroxymethyluracil (HmUra), which is repaired by HmUra-DNA glycosylase and does not block replication, will be compared to that of thymine glycol and related lesions which are repaired by E. coli endonuclease III and do block replication. HmUra-DNA glycosylase is unique among repair enzymes in being absent from bacteria and yeast. We have proposed that its phylogenetic distribution reflects its major function which is the repair of 5-methylcytosine residues in DNA. In contrast, endonuclease III-like activities are phylogenetically conserved indicating that the lesions repaired by the enzymes are formed in all species. Among these lesions are UV induced pyrimidine hydrates. The hydration of cytosine leads to irreversible deamination to uracil hydrate which, if unrepaired, leads to point mutations. This reaction scheme may also occur for 5-methylcytosine residues leading to irreversible deamination to thymine hydrate. We will study the formation and stability of pyrimidine photohydrates in oligonucleotides of high specific radioactivity using E. coli endonuclease III as a probe. We will use these oligonucleotides as substrates for the purification of mammalian endonuclease III-like activities to a purity sufficient to obtain protein sequence and molecular cloning. The results of these experiments will add to our knowledge of the formation and repair of oxidative and photochemical damage to DNA and possibly elucidate the relationship of such damage to the etiology of actinic and spontaneous cancer.