Project Details
Projekt
Molecular mechanisms of the interaction of chaotropic salts with natural and artificial DNA structures
Subject Area
Physical Chemistry of Molecules, Liquids and Interfaces, Biophysical Chemistry
Biophysics
Biophysics
Term
from 2019 to 2023
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 428230890
Due to their great versatility which enables the spatially selective immobilization of biomolecules with nanometer precision, DNA origami nanostructures are increasingly used substrates for the single-molecule investigation of various biochemical and biophysical processes. In these applications, the DNA origami substrates not only provide high positioning accuracy but also minimize thermal fluctuations due to their comparatively high mechanical rigidity. Consequently, the structural properties and especially the stability of the DNA origami substrates have become key issues in conducting such single-molecule studies. In this project, we will investigate the interaction of various chaotropic agents with different DNA structures at a molecular level. While chaotropic agents such as guanidinium (Gdm+) or tetrapropylammonium (TPA+) salts are established denaturants of protein structure and frequently used in protein unfolding studies, less is known about their interaction with DNA. Elucidating the mechanisms of their attack on DNA thus represents an important step toward DNA origami-based single-molecule studies of protein folding and unfolding dynamics. Molecular understanding of the processes involved in DNA origami denaturation by such agents, however, is further complicated by the various natural and non-natural DNA structures found within a single DNA origami. This project therefore aims at elucidating the mechanisms of chaotrope-DNA interactions with a particular focus on the role of DNA structure. This will be achieved by employing natural DNAs of different nucleotide sequence, simple artificial DNA nanostructures, and complex 2D and 3D DNA origami nanostructures. The latter enables the controlled induction of over- and underwound DNA duplex structures. Changes in the overall morphology of the DNA origami nanostructures and in particularly DNA origami denaturation will be monitored by atomic force microscopy while optical spectroscopy will yield detailed information about the molecular interactions of the chaotropes with the DNA helices. Using this experimental approach, counterion effects in the interaction of Gdm+ and TPA+ with DNA will be investigated that are known to play significant roles in protein denaturation. Furthermore, also the effect of hydrated cations will be addressed in order to elucidate the molecular mechanisms of salting-out-induced DNA denaturation.The experimental results of this project will thus provide a solid foundation for developing an understanding of the interaction of chaotropic agents such as Gdm+ and TPA+ with nucleic acids. By comparing genomic DNAs to artificial DNA nanostructures, also the so far only unsatisfyingly explored chemical peculiarities of non-natural DNA structures will be elucidated.
DFG Programme
Research Grants
