Binding-linked protonation of a DNA minor-groove agent
Binding-linked protonation of a DNA minor-groove agent
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DOI：
10.1529/biophysj.105.071381
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发表时间：
2006-02-01
影响因子：
3.4
通讯作者：
Wilson, WD
中科院分区：
文献类型：
--
作者：
Nguyen, B;Stanek, J;Wilson, WD
通过光谱学、等温滴定量热法和表面等离子体共振生物传感器方法研究了二苯基二脒抗锥虫药物 CGP 40215A 与 DNA 结合的能量学。两种脒在实验条件下都带正电，但两个苯基脒的连接基团的 pK(a) 为 6.3，容易发生质子化过程。光谱研究表明，当化合物与 A/T 小凹槽位点结合时，连接基团的 pKa 增加了 2.7 个单位。不同缓冲液和 pH 条件下的量热滴定支持质子连接过程，并且与光谱滴定非常一致。这两种方法建立了质子摄取亲和力。 le 作为 pH 值的函数。络合物形成的放热焓随不同pH条件而变化。观察到的结合热随着温度的变化而增加，表明负热容变化，这对于 DNA 小凹槽结合剂来说是典型的。溶剂可及表面积计算表明，表面埋藏约占观察到的内在负热容变化的一半。生物传感器和量热实验表明，由于质子化和静电相互作用，结合亲和力随 pH 值和盐浓度而变化。表面等离子体共振结合研究表明，DNA 发夹中每个磷酸盐的电荷密度小于聚合物中的电荷密度。还估计了配体/DNA 复合物的不同因素的能量贡献。
The energetics for binding of a diphenyl diamidine antitrypanosomal agent CGP 40215A to DNA have been studied by spectroscopy, isothermal titration calorimetry, and surface plasmon resonance biosensor methods. Both amidines are positively charged under experimental conditions, but the linking group for the two phenyl amidines has a pK(a) of 6.3 that is susceptible to a protonation process. Spectroscopic studies indicate an increase of 2.7 pKa units in the linking group when the compound binds to an A/T minor-groove site. Calorimetric titrations in different buffers and pH conditions support the proton-linkage process and are in a good agreement with spectroscopic titrations. The two methods established a proton-uptake pro. le as a function of pH. The exothermic enthalpy of complex formation varies with different pH conditions. The observed binding enthalpy increases as a function of temperature indicating a negative heat capacity change that is typical for DNA minor-groove binders. Solvent accessible surface area calculations suggest that surface burial accounts for about one-half of the observed intrinsic negative heat capacity change. Biosensor and calorimetric experiments indicate that the binding affinities vary with pH values and salt concentrations due to protonation and electrostatic interactions. The surface plasmon resonance binding studies indicate that the charge density per phosphate in DNA hairpins is smaller than that in polymers. Energetic contributions from different factors were also estimated for the ligand/DNA complex.