Mismatch Recognition by Saccharomyces cerevisiae Msh2-Msh6: Role of Structure and Dynamics

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• The mismatch repair (MMR) pathway is essential for maintaining genome integrity by correcting DNA replication errors.
• The Msh2-Msh6 protein complex is a key component of MMR, responsible for recognizing and initiating the repair of mismatches and small insertion/deletion loops.

Purpose of the Study:

• To investigate the role of local DNA dynamics in the recognition of mismatches by the Msh2-Msh6 complex.
• To determine how structural and dynamic elements of mismatches influence Msh2-Msh6 binding affinity and DNA bending.

Main Methods:

• Incorporation of the fluorescent nucleoside base analog 6-methylisoxanthopterin (6-MI) into DNA to probe mismatch properties.
• Fluorescence quantum yield and rotational correlation time measurements to assess local base dynamics and protein binding affinity.
• Förster Resonance Energy Transfer (FRET) measurements to quantify protein-induced DNA bending.

Main Results:

• A direct correlation was observed between local base dynamics and Msh2-Msh6 binding affinity, with higher affinity for more dynamic mismatches (K_D ≤ 25 nM).
• Protein-induced DNA bending was similar for both well- and poorly-recognized mismatches, suggesting bending is not the primary recognition determinant.
• Base-specific dynamics and spectral shifts support a directional binding model where Msh6's Phe 432 intercalates 3' of the mismatch.

Conclusions:

• Local base dynamics play a critical role in the initial recognition of DNA mismatches by the Msh2-Msh6 complex.
• The findings support a model of directional binding, influenced by the dynamic properties of the DNA mismatch.