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Mismatch repair proteins MutS and MutL inhibit RecA-catalyzed strand transfer between diverged DNAs
1Department of Biochemistry, Duke University Medical Center, Durham, NC 27710.
Abstract:
Bacterial mutS and mutL mutations confer large increases in recombination between sequences that are divergent by several percent at the nucleotide level, an effect attributed to a role for products of these genes in control of recombination fidelity. Since MutS and MutL are proteins involved in the earliest steps of mismatch repair, including mismatch recognition by MutS, we have tested the possibility that they may affect strand exchange in response to occurrence of mispairs within the recombination heteroduplex. We show that MutS abolishes RecA-catalyzed strand transfer between fd and M13 bacteriophage DNAs, which vary by 3% at the nucleotide level, but is without effect on M13-M13 or fd-fd exchange. Although MutL alone has no effect on M13-fd heteroduplex formation, the protein dramatically enhances the inhibition of strand transfer mediated by MutS. Analysis of strand-transfer intermediates that accumulate in the presence of MutS and MutL indicates that the proteins block branch migration, presumably in response to occurrence of mispairs within newly formed heteroduplex.
Insights
Bacterial MutS and MutL proteins regulate DNA recombination fidelity. These proteins inhibit strand exchange when DNA mismatches occur, preventing errors during genetic recombination.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- Bacterial mutS and mutL mutations increase recombination between divergent DNA sequences.
- MutS and MutL proteins are key players in DNA mismatch repair, involved in early recognition and processing of mismatches.
- Their role in controlling recombination fidelity suggests a potential influence on strand exchange processes.
Purpose of the Study:
- To investigate the role of bacterial MutS and MutL proteins in DNA strand exchange during recombination.
- To determine if MutS and MutL affect strand exchange in response to DNA sequence mispairs within a heteroduplex.
Main Methods:
- RecA-catalyzed strand transfer assays were performed using bacteriophage DNAs (fd and M13) with nucleotide sequence differences.
- The effect of purified MutS and MutL proteins, individually and in combination, on strand transfer was analyzed.
- Accumulation of strand-transfer intermediates was examined to understand the mechanism of inhibition.
Main Results:
- MutS protein abolished RecA-catalyzed strand transfer between divergent fd and M13 bacteriophage DNAs (3% nucleotide divergence).
- MutS had no effect on strand transfer between identical M13-M13 or fd-fd DNA molecules.
- MutL alone did not inhibit strand transfer, but significantly enhanced the inhibitory effect of MutS.
- Analysis of intermediates indicated that MutS and MutL block branch migration, likely in response to DNA mispairs.
Conclusions:
- Bacterial MutS and MutL proteins play a crucial role in suppressing recombination between divergent DNA sequences.
- These mismatch repair proteins inhibit strand exchange by blocking branch migration when DNA mispairs are present in the heteroduplex.
- This mechanism contributes to maintaining genetic recombination fidelity in bacteria.