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Application of Stopped-flow Kinetics Methods to Investigate the Mechanism of Action of a DNA Repair Protein
Published on: March 31, 2010
Affinity of mismatch-binding protein MutS for heteroduplexes containing different mismatches
1Division of Biochemistry and Molecular Biology, School of Biological Sciences, University of Southampton, Bassett Crescent East, Southampton S016 7PX, UK.
Abstract:
We have used bandshift analysis to measure the interaction between the Escherichia coli mismatch-binding protein MutS and synthetic DNA fragments containing all possible DNA mismatches as well as an unpaired T (DeltaT). The order of affinity is found to be DeltaT>GT>GG>AA approximately TT approximately TC>CA>GA>CC>GC. We find that the affinity for GT mismatches is affected by the flanking sequence and decreases in the order G(n)C(n)>(GC)(n)>A(n)T(n)>(AT)(n). Studies with base analogues show good binding to phiT (where phi represents 1',2'-dideoxyribose), but much weaker binding to Gphi.
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