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Real-time Observation of the DNA Strand Exchange Reaction Mediated by Rad51
Published on: February 13, 2019
Counting RAD51 proteins disassembling from nucleoprotein filaments under tension.
Joost van Mameren1, Mauro Modesti, Roland Kanaar
1Laser Centre and Department of Physics and Astronomy, VU University, De Boelelaan 1081, 1081 HV, Amsterdam, The Netherlands.
Nature
|December 9, 2008
Summary
Disassembly of RAD51 filaments, crucial for DNA repair, is driven by ATP hydrolysis and tension release. External tension on DNA stalls this process, revealing a tension-dependent mechanism at filament ends.
Area of Science:
- Molecular Biology
- Biochemistry
- Genetics
Background:
- Homologous recombination is a critical DNA repair pathway in eukaryotes.
- RAD51 protein filaments are central catalysts in this process, forming nucleoprotein filaments on single-stranded DNA.
- The disassembly mechanism of RAD51 filaments, essential for completing recombination, remains poorly understood.
Purpose of the Study:
- To elucidate the molecular mechanism of human RAD51 nucleoprotein filament disassembly.
- To investigate the roles of ATP hydrolysis and mechanical tension in RAD51 filament disassembly.
- To understand how accessory proteins might influence RAD51 filament dynamics.
Main Methods:
- Utilized a combination of optical tweezers, single-molecule fluorescence microscopy, and microfluidics.
- Applied external tension to DNA to study its effect on RAD51 filament disassembly.
- Quantified RAD51 protein fluorescence to monitor disassembly kinetics.
Main Results:
- RAD51 filament disassembly is a result of the interplay between ATP hydrolysis and the release of stored filament tension.
- Applying external tension to DNA significantly slows down or stalls RAD51 filament disassembly.
- Disassembly occurs in bursts interspersed with pauses, with tension-dependent disassembly occurring at filament ends after ATP hydrolysis.
Conclusions:
- The study reveals a tension-dependent mechanism for RAD51 filament disassembly, occurring at filament ends.
- ATP hydrolysis initiates disassembly, but tension release is required for its progression.
- This work provides insights into the regulation of homologous recombination by understanding RAD51 filament dynamics.
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