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Related Experiment Video

Updated: Jun 4, 2025

Detection of Homologous Recombination Intermediates via Proximity Ligation and Quantitative PCR in Saccharomyces cerevisiae

Detection of Homologous Recombination Intermediates via Proximity Ligation and Quantitative PCR in Saccharomyces cerevisiae

Published on: September 11, 2022

Structure guided functional analysis of the S. cerevisiae Mre11 complex.

John Petrini¹, Marcel Hohl², You Yu³

¹Memorial Sloan Kettering Cancer Center.

Research Square

|December 23, 2024

Summary

The Mre11-Rad50 complex structure reveals critical residues for DNA binding and assembly. This yeast complex is vital for DNA damage repair and signaling pathways.

Keywords:

Cryo-EM DNA repair Mre11 complex Rad50 coiled coil double strand break hook

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Last Updated: Jun 4, 2025

Detection of Homologous Recombination Intermediates via Proximity Ligation and Quantitative PCR in Saccharomyces cerevisiae

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Published on: September 11, 2022

Real-time Observation of the DNA Strand Exchange Reaction Mediated by Rad51

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Preparation of the Mgm101 Recombination Protein by MBP-based Tagging Strategy

Preparation of the Mgm101 Recombination Protein by MBP-based Tagging Strategy

Published on: June 25, 2013

Area of Science:

Molecular Biology
Structural Biology
Biochemistry

Background:

The Mre11 complex, comprising Mre11, Rad50, and Nbs1/Xrs2, is essential in eukaryotes for DNA double-strand break (DSB) repair.
Mre11 and Rad50 are highly conserved across life, while Nbs1/Xrs2 is eukaryotic-specific.
The complex plays a crucial role in detecting DNA damage and activating signaling pathways.

Purpose of the Study:

To determine the cryo-electron microscopy (cryo-EM) structure of the Saccharomyces cerevisiae Mre11-Rad50 complex bound to double-stranded DNA (dsDNA).
To elucidate the molecular mechanisms underlying complex assembly, DNA binding, and the functional roles of conserved residues.
To investigate the influence of the Rad50 coiled-coil domain on ATP hydrolysis.

Main Methods:

Cryo-electron microscopy (cryo-EM) to obtain a 3.2 Å resolution structure.
Biochemical assays and mutational analyses to investigate protein interfaces and DNA binding.
Structural analysis to identify conserved residues critical for complex function.

Main Results:

A high-resolution cryo-EM structure of the Mre11-Rad50-dsDNA complex was obtained.
Identified conserved residues in Mre11 and Rad50 essential for complex assembly and DNA interaction.
Demonstrated that the Rad50 coiled-coil domain allosterically affects ATP hydrolysis distant from the catalytic site.

Conclusions:

The determined structure provides a molecular basis for understanding Mre11-Rad50 complex function in DNA repair.
Conserved residues are key for the structural integrity and DNA-binding capabilities of the complex.
Rad50's coiled-coil domain plays a regulatory role in the complex's enzymatic activity.