Dipartimento di Scienze della Salute, Università di Milano-Bicocca, via Cadore 48, Monza (MB) 20900, Italy.
Nucleic Acids Res. 2013 Feb 1;41(3):2009-19. doi: 10.1093/nar/gks1206. Epub 2012 Dec 16.
The opening of DNA double strands is extremely relevant to several biological functions, such as replication and transcription or binding of specific proteins. Such opening phenomenon is particularly sensitive to the aqueous solvent conditions in which the DNA molecule is dispersed, as it can be observed by considering the classical dependence of DNA melting temperature on pH and salt concentration. In the present work, we report a single-molecule study of the stability of DNA against denaturation when subjected to changes in solvent. We investigated the appearance of DNA instability under specific external applied force and imposed twist values, which was revealed by an increase in the temporal fluctuations in the DNA extension. These fluctuations occur in the presence of a continuous interval of equilibrium states, ranging from a plectonemic state to a state characterized by denaturation bubbles. In particular, we observe the fluctuations only around a characteristic force value. Moreover, this characteristic force is demonstrated to be notably sensitive to variations in the pH and ionic strength. Finally, an extension of a theoretical model of plectoneme formation is used to estimate the average denaturation energy, which is found to be linearly correlated to the melting temperature of the DNA double strands.
DNA 双链的打开与许多生物学功能密切相关，例如复制和转录，或特定蛋白质的结合。这种打开现象对 DNA 分子分散在其中的水溶剂条件非常敏感，因为可以通过考虑 DNA 融解温度对 pH 值和盐浓度的经典依赖性来观察到这一点。在本工作中，我们报告了对 DNA 在溶剂变化下变性稳定性的单分子研究。我们研究了在特定外部施加力和规定的扭转值下 DNA 不稳定性的出现，这是通过 DNA 延伸的时间波动增加来揭示的。这些波动出现在从扭结态到以变性泡为特征的状态的连续平衡态范围内。特别是，我们仅在特征力值附近观察到波动。此外，该特征力被证明对 pH 值和离子强度的变化非常敏感。最后，使用扭结形成的理论模型的扩展来估计平均变性能，发现它与 DNA 双链的融解温度呈线性相关。
