REVERSE CHEMICAL MUTAGENESIS - IDENTIFICATION OF THE MUTAGENIC LESIONS RESULTING FROM REACTIVE OXYGEN SPECIES-MEDIATED DAMAGE TO DNA
REVERSE CHEMICAL MUTAGENESIS - IDENTIFICATION OF THE MUTAGENIC LESIONS RESULTING FROM REACTIVE OXYGEN SPECIES-MEDIATED DAMAGE TO DNA
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DOI：
10.1073/pnas.91.14.6609
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发表时间：
1994-07-05
影响因子：
11.1
通讯作者：
LOEB, LA
中科院分区：
文献类型：
--
作者：
FEIG, DI;SOWERS, LC;LOEB, LA
活性氧在DNA中引起的大量不同的化学修饰阻碍了对活性氧对诱变作用的理解。尽管这些DNA改变中的许多已经被编目，但导致突变的特定病变的鉴定仍然依赖于一次测试一种修饰。在这项研究中，我们提出了另一种方法，以确定关键的诱变性病变池的氧化修饰的核苷酸。用含有FeSO 4、H2 O2和抗坏血酸的氧自由基产生系统处理dCTP。通过反相和阴离子交换HPLC分离修饰产物，然后通过人免疫缺陷病毒逆转录酶将其掺入含有用于突变评分的靶基因的DNA中。分离的致突变物质之一被鉴定为5-羟基-2 '-脱氧胞苷。它被有效地整合到DNA中，并以2.5%的频率在大肠杆菌中引起C -> T转换，这比任何先前鉴定的氧化DNA损伤更具致突变性。
An understanding of the contribution of reactive oxygen species to mutagenesis has been hampered by the vast number of different chemical modifications they cause in DNA. Even though many of these DNA alterations have been catalogued, the identification of specific lesions that cause mutations has depended on testing one modification at a time. In this study we present another approach to identify key mutagenic lesions from a pool of oxidatively modified nucleotides. dCTP was treated with an oxygen radical-generating system containing FeSO4, H2O2, and ascorbic acid. The modification products were separated by reverse-phase and anion exchange HPLC and then incorporated by human immunodeficiency virus reverse transcriptase into a DNA that contains a target gene for scoring for mutations. One of the mutagenic species isolated was identified as 5-hydroxy-2'-deoxycytidine. It is incorporated efficiently into DNA and causes C --> T transitions in Escherichia coli at a frequency of 2.5%, which is more mutagenic than any previously identified oxidative DNA lesion.