Graziewicz M A, Zastawny T H, Oliński R, Tudek B
Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warsaw, Poland.
Mutat Res. 1999 May 14;434(1):41-52. doi: 10.1016/s0921-8777(99)00012-9.
Gas chromatography/isotope dilution-mass spectrometry with selected ion monitoring (GC/IDMS-SIM) was used to measure oxidised bases in hypoxanthine/xanthine oxidase/Fe3+/EDTA modified ss M13 mp18 phage DNA. A dose-dependent increase of oxidised bases content in DNA was observed with the biggest augmentation of FapyGua, thymine glycol and FapyAde. The amount of 8-OH-Gua was relatively high both in non-oxidised and oxidised DNA, and increased to the same extent as FapyAde and ThyGly. DNA oxidation caused a dramatic decrease in phage survival after transfection to E. coli. Survival was improved 2.8-fold after induction of the SOS system by UV irradiation of bacteria and mutation frequency of the lacZ gene in SOS conditions increased 7-fold over that in non-irradiated bacteria. Spectrum of mutations was different from those reported previously and mutations were distributed rather randomly within M13 lacZ sequence, which was in contrast to previous findings, where with non-chelated metal ions other types of mutations were found in several clusters. Thus, conditions of DNA oxidation and accessibility of metal ions for DNA bases might be important factors for generating different DNA damages and mutations. Major base substitutions found both in SOS-induced and non-induced E. coli but with higher mutation frequency in SOS-induced cells were C-->A (approximately 20-fold increase in SOS-conditions), G-->A (9-fold increase) and G-->C (4.5-fold increase). Very few G-->T transitions were found. A particularly large group of A-->G transitions appeared only in SOS-induced bacteria and was accompanied by augmentation of FapyAde content in the phage DNA with undetectable 2-OH-Ade. It is then possible that imidazole ring-opened adenine mimics guanine during DNA replication and pairs with cytosine yielding A-->G transitions in SOS-induced bacteria.
采用气相色谱/同位素稀释-质谱联用选择离子监测法（GC/IDMS-SIM）测定次黄嘌呤/黄嘌呤氧化酶/Fe3+/乙二胺四乙酸（EDTA）修饰的单链M13 mp18噬菌体DNA中的氧化碱基。观察到DNA中氧化碱基含量呈剂量依赖性增加，其中FapyGua、胸腺嘧啶乙二醇和FapyAde增加最为显著。8-羟基鸟嘌呤（8-OH-Gua）在未氧化和氧化的DNA中含量都相对较高，且与FapyAde和胸腺嘧啶乙二醇（ThyGly）的增加幅度相同。DNA氧化导致转染大肠杆菌后噬菌体存活率显著下降。在用紫外线照射细菌诱导SOS系统后，存活率提高了2.8倍，并且在SOS条件下，lacZ基因的突变频率比未照射细菌时增加了7倍。突变谱与先前报道的不同，突变在M13 lacZ序列中分布较为随机，这与先前的研究结果相反，先前研究发现，在使用非螯合金属离子时，会在几个簇中发现其他类型的突变。因此，DNA氧化条件以及金属离子与DNA碱基的可及性可能是产生不同DNA损伤和突变的重要因素。在SOS诱导和未诱导的大肠杆菌中均发现了主要的碱基替换，但在SOS诱导的细胞中突变频率更高，包括C→A（在SOS条件下增加约20倍）、G→A（增加9倍）和G→C（增加4.5倍）。发现的G→T转换很少。特别大的一组A→G转换仅出现在SOS诱导的细菌中，并且伴随着噬菌体DNA中FapyAde含量的增加，而未检测到2-羟基腺嘌呤（2-OH-Ade）。那么有可能在DNA复制过程中，咪唑环打开的腺嘌呤模拟鸟嘌呤并与胞嘧啶配对，从而在SOS诱导的细菌中产生A→G转换。
