Calorimetric and spectroscopic studies of Hoechst 33258: Self-association and binding to non-cognate DNA
Calorimetric and spectroscopic studies of Hoechst 33258: Self-association and binding to non-cognate DNA
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DOI：
10.1016/j.jmb.2008.05.073
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发表时间：
2008-09-05
影响因子：
5.6
通讯作者：
Haq, Ihtshamul
中科院分区：
文献类型：
--
作者：
Buurma, Niklaas J.;Haq, Ihtshamul
小分子DNA的序列和结构特异性分子识别是生物物理化学和药物发现的重要目标。许多候选配体具有平坦的芳香表面和其他分子特征，使它们能够自结合。此外，与靶标的非特异性结合是这些相互作用的一个复杂特征。因此，为了获得有意义的热力学参数，需要在数据分析中考虑多重平衡。为了解决这些问题，我们系统地研究了双苯并咪唑染料Hoechst 33258 （H33258）的自聚集和结合DNA寡核苷酸缺乏任何同源的小凹槽A-T位点。用等温滴定量热法（ITC）、圆二色法（CD）、荧光光谱法和脉冲梯度自旋回波核磁共振（NMR）对该模型进行了研究。已经确定了三种不同的结合事件和配体自聚集，并在可能的情况下进行了量化。H33258自聚集涉及由堆叠相互作用驱动的逐步聚集机制。DNA结合过程包括两种特异性结合模式和非特异性DNA模板H33258叠加模式。我们编写了新颖的ITC数据拟合软件（IC-ITC；免费提供给生物物理学界），它同时适合配体聚集和配体- dna结合。在这里，这个数值分析，它使用模拟退火的复杂量热数据表示多个耦合平衡，是描述。(C) 2008 Elsevier Ltd版权所有。
Sequence and structure-specific molecular recognition of DNA by small molecules is an important goal in biophysical chemistry and drug discovery. Many candidate ligands possess flat aromatic surfaces and other molecular features that allow them to self-associate. In addition, non-specific binding to the target is a complicating feature of these interactions. Therefore, multiple equilibria are present and need to be accounted for in data analysis in order to obtain meaningful thermodynamic parameters. In order to address these issues we have systematically examined the bis-benzimidazole dye Hoechst 33258 (H33258) in terms of self-aggregation and binding to DNA oligonucleotides lacking any cognate minor groove A-T sites. This model system has been interrogated using isothermal titration calorimetry (ITC), circular dichroism (CD), fluorescence spectroscopy and pulsed gradient spin echo NMR. Three distinct binding events and ligand self-aggregation have been identified and, where possible, quantified. H33258 self-aggregation involves a step-wise aggregation mechanism, driven by stacking interactions. The DNA binding process includes two specific binding modes and non-specific DNA-templated H33258 stacking. We have written novel ITC data-fitting software (IC-ITC; freely available to the biophysics community), which simultaneously fits ligand aggregation and ligand-DNA binding. Here, this numerical analysis, which uses simulated annealing of complex calorimetric data representing multiple coupled equilibria, is described. (C) 2008 Elsevier Ltd. All rights reserved.