Department of Chemistry, State Key Laboratory of Coordination Chemistry, Nanjing University, Nanjing 210093, Jiangsu, PR China.
Org Biomol Chem. 2012 Nov 14;10(42):8484-92. doi: 10.1039/c2ob25743b. Epub 2012 Sep 25.
A new metal-free DNA cleaving reagent, bis-tacnorthoamide derivative 1 with two tacnorthoamide (tacnoa) units linked by a spacer containing anthraquinone, has been synthesized from triazatricyclo[5.2.1.0(4,10)]decane and characterized by NMR and mass spectrometry. For comparison, the corresponding compounds mono-tacnorthoamide derivative 2 with one tacnorthoamide unit and 6 with two tacnorthoamide units linked by an alkyl (1,6-hexamethylene) spacer without anthraquinone have also been synthesized. The DNA-binding property investigated via fluorescence and CD spectroscopy suggests that compounds 1 and 2 have an intercalating DNA binding mode, and the apparent binding constants of 1, 2 and 6 are 1.3 × 10(7) M(-1), 0.8 × 10(7) M(-1) and 8 × 10(5) M(-1), respectively. Agarose gel electrophoresis was used to assess plasmid pUC19 DNA cleavage activity promoted by 1, 2, 6 and parent tacnoa under physiological conditions, which gives rate constants k(obs) of 0.2126 ± 0.0055 h(-1), 0.0620 ± 0.0024 h(-1), 0.040 ± 0.0007 h(-1) and 0.0043 ± 0.0002 h(-1), respectively. The 50-fold and 15-fold rate acceleration over parent tacnoa is because of the anthraquinone moiety of compound 1 or 2 intercalating into DNA base pairs via a stacking interaction. Moreover, DNA cleavage reactions promoted by compound 1 give 5.3-fold rate acceleration over compound 6, which further demonstrates that the introduction of anthraquinone results in a large enhancement of DNA cleavage activity. In particular, DNA cleavage activity promoted by 1 bearing two tacnoa units is 3.3 times more effective than 2 bearing one tacnoa unit and the DNA cleavage by compound 1 was achieved effectively at a relatively low concentration (0.03 mM). This dramatic rate acceleration suggests the cooperative catalysis of the two positively charged tacnoa units in compound 1. The radical scavenger inhibition study and ESI-MS analysis of bis(2,4-dinitrophenyl) phosphate (BDNPP) and adenylyl(3'-5')phosphoadenine (APA) cleavage in the presence of compound 1 suggest the cleavage mechanism would be via a hydrolysis pathway by cleaving the phosphodiester bond of DNA.
一种新型的无金属 DNA 切割试剂，双 tacnorthoamide 衍生物 1，由三氮杂三环[5.2.1.0(4,10)]癸烷和含有蒽醌的间隔物连接两个 tacnorthoamide(tacnoa)单元合成，并通过 NMR 和质谱进行了表征。为了进行比较，还合成了相应的化合物单 tacnorthoamide 衍生物 2，具有一个 tacnorthoamide 单元，和 6，通过无蒽醌的烷基(1,6-己二甲基)间隔物连接两个 tacnorthoamide 单元。通过荧光和 CD 光谱研究 DNA 结合特性表明，化合物 1 和 2 具有嵌入 DNA 的结合模式，并且 1、2 和 6 的表观结合常数分别为 1.3×10(7)M(-1)、0.8×10(7)M(-1)和 8×10(5)M(-1)。琼脂糖凝胶电泳用于评估生理条件下 1、2、6 和母体 tacnoa 对质粒 pUC19 DNA 的切割活性，得到 1 的观察速率常数 k(obs)为 0.2126±0.0055h(-1)，2 为 0.0620±0.0024h(-1)，6 为 0.040±0.0007h(-1)，母体 tacnoa 为 0.0043±0.0002h(-1)。由于化合物 1 或 2 的蒽醌部分通过堆积相互作用嵌入 DNA 碱基对，与母体 tacnoa 相比，50 倍和 15 倍的速率加速。此外，化合物 1 促进的 DNA 切割反应比化合物 6 快 5.3 倍，这进一步证明引入蒽醌可大大增强 DNA 切割活性。特别是，具有两个 tacnoa 单元的化合物 1 的 DNA 切割活性比具有一个 tacnoa 单元的 2 高 3.3 倍，并且化合物 1 在相对较低的浓度(0.03 mM)下有效实现 DNA 切割。这种显著的速率加速表明化合物 1 中的两个带正电荷的 tacnoa 单元具有协同催化作用。在存在化合物 1 的情况下，通过自由基清除剂抑制研究和二(2,4-二硝基苯基)磷酸酯(BDNPP)和腺嘌呤基(3'-5')磷酸腺嘌呤(APA)的 ESI-MS 分析，表明切割机制将通过切割 DNA 的磷酸二酯键而通过水解途径进行。
