Reis L A, Rocha M S
Laboratório de Física Biológica, Departamento de Física, Universidade Federal de Viçosa, Minas Gerais, Brazil.
Biopolymers. 2017 May;107(5). doi: 10.1002/bip.23015.
In this work, we use force spectroscopy to investigate the interaction between the DAPI fluorescent dye and the λ-DNA molecule under high (174 mM) and low (34 mM) ionic strengths. Firstly, we have measured the changes on the mechanical properties (persistence and contour lengths) of the DNA-DAPI complexes as a function of the dye concentration in the sample. Then, we use recently developed models in order to connect the behavior of both mechanical properties to the physical chemistry of the interaction. Such analysis has allowed us to identify and to decouple two main binding modes, determining the relevant physicochemical (binding) parameters for each of these modes: minor groove binding, which saturates at very low DAPI concentrations ( CT ∼ 0.50 μM) and presents equilibrium binding constants of the order of ∼10 M for the two ionic strengths studied; and intercalation, which starts to play a significant role only after the saturation of the first mode, presenting much smaller equilibrium binding constants (∼10 M ).
在这项工作中，我们使用力谱法研究了在高（174 mM）和低（34 mM）离子强度下，DAPI荧光染料与λ-DNA分子之间的相互作用。首先，我们测量了DNA-DAPI复合物的机械性能（持久长度和轮廓长度）随样品中染料浓度的变化。然后，我们使用最近开发的模型，以便将这两种机械性能的行为与相互作用的物理化学联系起来。这种分析使我们能够识别并解耦两种主要的结合模式，确定每种模式的相关物理化学（结合）参数：小沟结合，在非常低的DAPI浓度（CT ∼ 0.50 μM）下达到饱和，对于所研究的两种离子强度，其平衡结合常数约为10 M；以及嵌入，仅在第一种模式饱和后才开始起重要作用，其平衡结合常数要小得多（∼10 M）。
