Computational studies of the mechanism of DNA strand-exchange reaction

NAISS 2023/5-545


NAISS Medium Compute

Principal Investigator:

Bengt Nordén


Chalmers tekniska högskola

Start Date:


End Date:


Primary Classification:

10603: Biophysics

Secondary Classification:

10602: Biochemistry and Molecular Biology

Tertiary Classification:

10407: Theoretical Chemistry



Preservation of genetic information over many generations requires high-fidelity of DNA homologous recombination reaction, which involves the genetic information exchange between DNA molecules. This biological process allows genetic diversity and is vital for cell survival by error-free recombinational repair of double-strand breaks, the most severe kind of DNA damage. The reaction is catalysed by RecA and Rad51 proteins in prokaryotes and eukaryotes, respectively. The enzymes bind a single-stranded part of DNA to form a filamentous complex, where the protein monomers are organised in a helical manner around DNA, which is stretched and underwound. The single-stranded DNA-RecA/Rad51 filament then interacts with a double-stranded DNA to form a ssDNA-RecA/Rad51-dsDNA complex. When and if the two DNA molecules are homologous, strand exchange occurs. Despite the importance of recombinases in many medicinal contexts (e.g. cancer, gene therapy, sterility) and years of intense research, the mechanistic details of the search for the sequential homology and the strand exchange are poorly understood at atomic level and many questions, including why DNA inside is stretched, remain enigmatic. One of the reasons why the recombination mechanisms have remained unsolved is that we lack understanding the DNA interactions in hydrophobic environments, created inside the RecA/Rad51 filaments. Recent evidence from spectroscopic data and optical tweezer experiments points towards an unexplored hydrophobic mechanism. Certain semi-hydrophobic co-solutes change the nucleobase stacking, leading to increased DNA flexibility, transient unstacking events, and lowered activation energy to spontaneous strand exchange between homologous DNA seqeunces. The determination of the RecA-ssDNA and RecA-dsDNA crystal structures revealed that the stretched DNA conformation has near perpendicular nucleobases orientation and clustering of bases triplets stacked approximately as in B-form DNA. The triplet arrangement may potentially have a biological role in enhancing the recognition of complementary bases sequence and promote the strand-exchange process. In the project we will study DNA stretching and consequently DNA strand-exchange mechanisms in varied hydrophobic environments, including dimethyl ethylene glycol, diglyme, diethyl ethylene glycol and 1,4-dioxane. In contrast to previous studies, where the DNA deformation is achieved via controlling boundary conditions or by restricting helical parameters, which ignores DNA sequence effects, we will use extended atomic MD simulations combined with the new DNA stretching restraint, designed to control the helical rise (axial extension) of a nucleic acid fragment without restraining any other helical parameter. Our approach describes DNA deformation as it happens in cells, as it allows molecules to freely explore the conformational space. We will also study the role of DNA sequence-composition, by testing several DNA sequences. We will perform MD simulations for the systems that includes a stretched dsDNA fragment and a homologous ssDNA fragment in a hydrophobic environment, resembling the one inside the RecA/Rad51 filaments to explore the sequence-specific effects and role of hydrophobic environment on DNA strand-exchange reaction. We expect that the gained mechanistic insights will contribute greatly to our understanding of homologous recombination reaction.