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Updated: Jun 6, 2025
06:51
Parallel High Throughput Single Molecule Kinetic Assay for Site-Specific DNA Cleavage
Published on: May 6, 2020
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Loop-extrusion-mediated plasmid DNA cleavage by the bacterial SMC Wadjet complex
Biswajit Pradhan1, Amar Deep2, Jessica König1
1Max Planck Institute of Biophysics, 60438 Frankfurt am Main, Germany.
Molecular Cell
|December 3, 2024
Summary
Structural maintenance of chromosomes (SMC) complexes like Wadjet use DNA loop extrusion to recognize and cleave plasmid DNA, defending against transformation. This mechanism is conserved across life.
Area of Science:
- Molecular Biology
- Genetics
- Microbiology
Background:
- Structural maintenance of chromosomes (SMC) complexes are crucial for genome organization and maintenance in all life forms.
- Prokaryotic Wadjet complexes, similar to MukBEF, prevent plasmid transformation by cleaving plasmid DNA, but the mechanism is unknown.
Purpose of the Study:
- To elucidate the molecular mechanism of plasmid recognition and cleavage by the Wadjet complex.
- To visualize the DNA processing activities of Wadjet using single-molecule imaging.
Main Methods:
- In vitro single-molecule imaging techniques were employed.
- Surface-anchored plasmid DNAs were used to observe Wadjet's interaction with DNA.
Main Results:
- Wadjet functions as a symmetric DNA loop extruder, requiring a dimeric JetABC supercomplex.
- Wadjet extrudes the entire length of a 44-kb plasmid, stalls, and subsequently cleaves the DNA.
- This loop extrusion activity is essential for specific plasmid recognition and elimination.
Conclusions:
- DNA loop extrusion is the mechanism by which Wadjet specifically recognizes and eliminates plasmids.
- Loop extrusion is an evolutionarily conserved mechanism among SMC complexes across all kingdoms of life.
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