Influence of the amino substituents in the interaction of ethidium bromide with DNA
Influence of the amino substituents in the interaction of ethidium bromide with DNA
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DOI：
10.1529/biophysj.104.047415
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发表时间：
2004-12-01
影响因子：
3.4
通讯作者：
Graves, DE
中科院分区：
文献类型：
--
作者：
Garbett, NC;Hammond, NB;Graves, DE
合理设计新型 DNA 结合剂的关键步骤是获得小分子-DNA 相互作用的完整热力学表征。四十多年来，溴化乙锭一直作为经典的 DNA 嵌入剂。这项工作的重点是描述乙锭的 3- 和 8- 氨基取代基对 DNA 复合物形成的能量贡献和伴随荧光特性的影响。去除 3- 或 8- 氨基取代基后，结合亲和力会降低一个数量级，而在两个氨基都不存在的情况下，结合亲和力会进一步降低一个数量级。当两个氨基都被去除时，热力学结合机制从母体乙锭的焓驱动变为熵驱动。 DNA 结合后，在存在一个或两个氨基的情况下观察到荧光增强，这可能是因为与埋入嵌入位点时相比，通过游离氨基的溶剂相互作用进行更有效的荧光猝灭。脱氨基乙锭类似物在结合时表现出荧光猝灭，这与通过与溶剂相互作用的发色团猝灭效率低于嵌入位点内的猝灭效率一致。量子效率的测定表明 DNA 结合位点内 3- 和 8- 氨基取代基的环境存在明显差异。
A key step in the rational design of new DNA binding agents is to obtain a complete thermodynamic characterization of small molecule-DNA interactions. Ethidium bromide has served as a classic DNA intercalator for more than four decades. This work focuses on delineating the influence(s) of the 3- and 8-amino substituents of ethidium on the energetic contributions and concomitant fluorescent properties upon DNA complex formation. Binding affinities decrease by an order of magnitude upon the removal of either the 3- or 8-amino substituent, with a further order-of-magnitude decrease in the absence of both amino groups. The thermodynamic binding mechanism changes from enthalpy-driven for the parent ethidium to entropy-driven when both amino groups are removed. Upon DNA binding, fluorescence enhancement is observed in the presence of either or both of the amino groups, likely because of more efficient fluorescence quenching through solvent interactions of free amino groups than when buried within the intercalation site. The des-amino ethidium analog exhibits fluorescence quenching upon binding, consistent with less efficient quenching of the chromophore through interactions with solvent than within the intercalation site. Determination of the quantum efficiencies suggests distinct differences in the environments of the 3- and 8-amino substituents within the DNA binding site.