MUTAGENIC CONSEQUENCES OF DEFINED LESIONS IN DNA

将确定的损伤引入DNA的致突变后果是 在一个突变系统中进行检测， 突变事件，无论是在实际的网站和一些距离， 的损害。 该系统已首次应用于脱嘌呤， DNA中的嘌呤碱基 这是一种常见的自发性病变， 在许多其他类型的DNA损伤修复中的常见中间体。 脱尿具有高度致突变性，通过检测脱嘌呤 MI3mp2 DNA导入SOS诱导的感受态E. coli细胞。 DNA序列分析 211个突变体中，大多数突变是碱基替换 反映了dAMP在非编码脱碱基位点对面的插入， 导致特征性的颠换。 这种颠换是 在体内经常观察到的几种损伤剂， DNA加合物，表明这种加合物可能产生突变， 通过无碱基位点中间体，或通过简单地排除模板碱基 氢键 第二个确定的损伤是DNA中的尿嘧啶残基，由DNA中的DNA片段引起。 热诱导胞嘧啶脱氨基。 这被认为是第二个 DNA中最常见的自发性病变，代表了一个显著的 在37摄氏度下挑战细胞。 对于这些研究，第一个体外研究 已经开发了脯氨酸密码子（CCC）回复的测定。 测定 是高度特异性和敏感性的 用这个和 正向突变试验中，C脱氨基的速率常数已被 在生理pH和盐浓度和337摄氏度下测定。 C脱氨基作用的序列特异性刚刚受到检验。 最终，各种加合物的交叉链质子化的贡献 在双链DNA中，导致C脱氨基，将被研究。

The mutagenic consequences of introducing defined lesions into DNA are being examined in a mutational system capable of detecting a wide spectrum of mutational events, both at and some distance away from the actual site of damage. The system has first been applied to depurination, the loss of a purine base from DNA. This is a frequent spontaneous lesion as well as a common intermediate in the repair of many other types of DNA damage. Depurination is highly mutagenic, as determined by transfecting depurinated MI3mp2 DNA into SOS-induced competent E. coli cells. DNA sequence analysis of 211 mutants demonstrates that most mutations are base substitutions reflecting insertion of dAMP opposite the non-coding abasic site and resulting in characteristic transversions. Such transversions are frequently observed in vivo for several damaging agents which produce bulky DNA adducts, suggesting that such adducts may produce mutations either through an abasic site intermediate, or by simply precluding template base hydrogen bonding. The second defined lesion is a uracil residue in DNA, resulting from the heat-induced deamination of cytosine. This is suggested to be the second most frequent spontaneous lesion in DNA, representing a significant challenge to a cell at 37 degrees C. For these studies, the first in vitro assay for reversion of a proline codon (CCC) has been developed. The assay is both highly specific and very sensitive. Using this and the forward-mutation assay, the rate constant of C deamination has been determined at physiological pH and salt concentrations and 3 37 degrees C. The sequence specificity of C deamination has just come under examination. Eventually, the contribution of cross-strand protonation by various adducts in double-stranded DNA, leading to C deamination, will be investigated.