The in vitro methylation of DNA by a minor groove binding methyl sulfonate ester
The in vitro methylation of DNA by a minor groove binding methyl sulfonate ester
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DOI：
10.1021/tx9501849
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发表时间：
1996-04-01
影响因子：
4.1
通讯作者：
Gold, B
中科院分区：
文献类型：
--
作者：
Encell, L;Shuker, DEG;Gold, B
基于附加有 O-甲基磺酸酯官能团的 N-甲基吡咯甲酰胺亚基 (MeOSO(2)(CH2)(2)-Lex) 的序列和沟槽特异性 DNA 甲基化剂的制备先前已有描述 [Zhang, Y., Chen, F.-X., Mehta, P., and Gold, B. (1993) Biochemistry 32, 7954-7965]。与主要在 7-鸟嘌呤甲基化的简单甲基磺酸酯（例如甲磺酸甲酯 (MMS)）相比，MeOSO(2)(CH2)(2)-Lex 提供 N3-甲基腺嘌呤 (3-MeAde) 作为其主要加合物。使用竞争性 ELISA 测定，可以精确定量 MeOSO(2)(CH2)(2)-Lex 对小牛胸腺 DNA 大沟和小沟位点的甲基化。使用 10 μM MeOSO(2)(CH2)(2)-Lex 和 100 μM DNA，N7-甲基鸟嘌呤 (7-MeGua)、3-MeAde 和 O-6-甲基脱氧鸟苷 (B-Me-dGuo) 的产量分别为 0.424、3.195 和 0.0027 mmol 加合物/mol DNA。相比之下，使用 MMS 时，7-MeGua、S-MeAde 和 6-Me-dGuo 的加合物/mol DNA 分别为 0.773、0.072 和 0.0033 mmol 加合物。由于 MeOSO(2)(CH2)(2)-Lex 的小沟平衡结合特性，3-MeAde 的产率增加，与 MMS 相比增加了 40 倍。
The preparation of sequence and groove specific DNA methylating agents based on N-methylpyrrolecarboxamide subunits appended with an O-methyl sulfonate ester functionality (MeOSO(2)(CH2)(2)-Lex) has previously been described [Zhang, Y., Chen, F.-X., Mehta, P., and Gold, B. (1993) Biochemistry 32, 7954-7965]. In contrast to simple methyl sulfonate esters, e.g., methyl methanesulfonate (MMS), which predominantly methylate at 7-guanine, MeOSO(2)(CH2)(2)-Lex affords N3-methyladenine (3-MeAde) as its major adduct. Using competitive ELISA determinations, the methylation at major and minor groove sites in calf thymus DNA by MeOSO(2)(CH2)(2)-Lex has been precisely quantitated. The yields of N7-methylguanine (7-MeGua), 3-MeAde, and O-6-methyldeoxyguanosine (B-Me-dGuo) are 0.424, 3.195, and 0.0027 mmol of adduct/mol of DNA, respectively, using 10 mu M MeOSO(2)(CH2)(2)-Lex and 100 mu M DNA. This compares to 0.773, 0.072, and 0.0033 mmol of adduct/mol of DNA for 7-MeGua, S-MeAde, and 6-Me-dGuo, respectively, using MMS. The increase in the yield of 3-MeAde due to the minor groove equilibrium binding properties of MeOSO(2)(CH2)(2)-Lex is similar to 40-fold relative to MMS.