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Characterization of a DNA mismatch-binding activity in yeast extracts
J J Miret1, M G Milla, R S Lahue
1Department of Biochemistry and Molecular Biology, University of Massachusetts Medical Center, Worcester 01655.
The Journal of Biological Chemistry
|February 15, 1993
Summary
A DNA mismatch binding activity in yeast recognizes various DNA mismatches, with MSH2 protein being crucial for this function. This finding is important for understanding DNA repair mechanisms.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- DNA replication fidelity is maintained by DNA mismatch repair (MMR) pathways.
- MMR corrects errors that escape proofreading during DNA replication.
Purpose of the Study:
- To identify and characterize DNA mismatch binding activities in yeast Saccharomyces cerevisiae.
- To investigate the role of MSH2 and MSH3 proteins in DNA mismatch recognition.
Main Methods:
- Band shift assays were used to detect DNA mismatch binding activity.
- Competition experiments were performed to assess binding specificity.
- UV cross-linking was employed to estimate protein molecular weight.
- Analysis of extracts from msh2 and msh3 mutant strains.
Main Results:
- A nuclear extract activity specifically bound to DNA oligonucleotides containing mismatches.
- The binding activity recognized various mispairs, including single base mismatches and deletions.
- Binding was dependent on the MSH2 protein, as it was absent in msh2- strains.
- The major binding protein had an estimated molecular weight of approximately 110 kDa, consistent with Msh2.
Conclusions:
- Yeast possesses a DNA mismatch binding activity crucial for DNA repair.
- MSH2 is essential for this mismatch binding activity.
- This activity likely plays a significant role in the Saccharomyces cerevisiae DNA mismatch repair pathway.