Magnetic tweezers measurements of the nanomechanical stability of DNA against denaturation at various conditions of pH and ionic strength

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 obs...

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Bibliographic Details
Published in:Nucleic acids research 2013-02, Vol.41 (3), p.2009-2019
Main Authors: Tempestini, Alessia, Cassina, Valeria, Brogioli, Doriano, Ziano, Roberto, Erba, Simona, Giovannoni, Roberto, Cerrito, Maria G, Salerno, Domenico, Mantegazza, Francesco
Format: Article
Language:English
Subjects:
Biomechanical Phenomena
DNA - chemistry
Hydrogen-Ion Concentration
Nanotechnology
Nucleic Acid Denaturation
Osmolar Concentration
Structural Biology
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Summary: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.
ISSN:0305-1048
1362-4962
DOI:10.1093/nar/gks1206