UV-induced photoproducts of 5-methylcytosine in a DNA sequence context.
UV-induced photoproducts of 5-methylcytosine in a DNA sequence context.
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DNA 序列背景下紫外线诱导的 5-甲基胞嘧啶光产物。
DOI：
10.1093/nar/16.8.3327
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发表时间：
1988
影响因子：
14.9
通讯作者：
Lapeyre,JN
中科院分区：
文献类型：
--
作者：
Barna,T;Malinowski,J;Holton,P;Ruchirawat,M;Becker,FF;Lapeyre,JN
为了检测可能的m5 C光产物，高度纯化的大鼠肝DNA-胞嘧啶甲基转移酶被用来特异性地产生具有放射性甲基的m5 C。当这些DNA受到大剂量（10 kJ/m2）的254 nm或302 nm紫外光（UVB）照射以提高产率时，经甲酸水解后，通过反相HPLC检测并分离出两种标记的光产物。使用丙酮作为三重态敏化剂和UVB照射的进一步研究表明，光产物II通过三重态被激活，而更极性的光产物I则没有。用10 kJ/m2254 nm的光对纯化的光产物进行光致逆转，证明了以下反应：光产物I再生m5 C，而光产物II被分裂并再生nrC和光产物I。这些结果表明，光产物I是单体，而光产物II二聚体，并从后者的洗脱位置可能是一个环丁基型二聚体所产生的反应与相邻的胞嘧啶。使用d[TTG]和d[Cm 5CG]作为典型序列的模型，分别用10 kJ/m2254 nm或302 nm照射，产生了在测序凝胶中具有改变的迁移率的小组分。改变的流动性三核苷酸耐降解PI和微球菌核酸酶从嘧啶碱基的光二聚化的预期。此外，含有m5 C的寡核苷酸底物的合成和显示是敏感的T4核酸内切酶V行动在位置一致的d[Cm 5C]光二聚体形成时，在UVB范围内照射。
In order to detect possible m5C photoproducts, highly purified rat liver DNA-cytos1ne methyltransferase was used to specifically generate m5C with a radioactive methyl group. When these DNAs were subjected to a large dose (10 kJ/m2) of 254 nm or 302 nm ultraviolet light (UVB) to enhance the yield, two labeled photoproducts were detected and isolated by reverse phase HPLC after formic add hydrolysis. Further studies using acetone as a triplet state sensitizer and UVB irradiation suggested that photoproduct II was activated via a triplet state while the more polar photoproduct I was not. Photoreversion of the purified photoproducts with 10 kJ/m2254 nm light demonstrated the following reactions: Photoproduct I regenerated m5C, while photoproduct II is split and regenerated nrC and photoproduct I. These results suggest that photoproduct I is monomeric while photoproduct II dimeric, and from the latter's elution position possibly a cyclobutyl type dimer arising from a reaction with an adjacent cytosine. Using d[TTG] and d[Cm5CG] as models of typical sequences, irradiation with 10 kJ/m2254 nm or 302 nm, respectively, gave rise to a small component having altered mobility in sequencing gels. The altered mobility trinucleotides were resistant to degradation by PI and micrococcal nucleases as expected from photodimerization of the pyrimidine bases. Furthermore, oligonucleotide substrates containing m5C were synthesized and shown to be susceptible to T4 endonuclease v action at locations consistent with d[Cm5C] photodimer formation when irradiated in the UVB range.