Saccharomyces cerevisiae Xrs2 Binds DNA Through Its FHA Domain

Area of Science:

• Molecular Biology
• Biochemistry
• Structural Biology

Background:

• The MRE11-RAD50-NBS1/Xrs2 (MRN/X) complex is crucial for responding to DNA double-strand breaks (DSBs).
• While DNA binding sites in MRE11 and RAD50 are known, the interaction of NBS1/Xrs2 with DNA remains structurally uncharacterized.

Purpose of the Study:

• To identify and structurally characterize the DNA binding site within the NBS1/Xrs2 component of the MRN/X complex.
• To elucidate the structural basis for Xrs2's interaction with DNA and its potential role in DSB repair regulation.

Main Methods:

• Nuclear Magnetic Resonance (NMR) spectroscopy (chemical shift perturbations and paramagnetic relaxation enhancements) to map DNA binding.
• Integrative modeling using HADDOCK to generate structural models of DNA-bound Xrs2.
• Site-directed mutagenesis and comparative binding assays to validate interaction interfaces.

Main Results:

• A direct DNA binding site was identified within the N-terminal FHA domain of Saccharomyces cerevisiae Xrs2.
• DNA binding at this site overlaps with the binding region for phosphorylated Sae2 peptide.
• NMR relaxation data indicate altered protein dynamics upon DNA binding, without changes in slower conformational exchange.

Conclusions:

• This study defines a direct DNA binding role for Xrs2, a critical component of the MRN/X complex.
• The findings provide a structural basis for Xrs2's dual recognition of DNA and phosphoproteins during DNA double-strand break repair.
• The results highlight the importance of integrating structural and functional studies for validating computational models.