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Chapters
- 00:00Title
- 00:57Introduction
- 01:40Sample Preparation for the Msh2-Msh6/ DNA Binding Experiment
- 03:07Instrument Preparation for the Msh-2Msh6/DNA Binding Experiment
- 05:07Msh2-Msh6/DNA Binding Experiment
- 06:27Representative Results for the Msh2-Msh6/DNA Binding Kinietics Experiment
- 07:00Sample and Instrument Preparation for the Msh2-Msh6 ATPase Experiment
- 08:30Msh2-Msh6 ATPase Experiment
- 09:40Representative Results for Msh2-Msh6 ATPase Kinetics Experiments
- 10:20Conclusion
Msh2-Msh6 is responsible for initiating repair of replication errors in DNA. Here we present a transient kinetics approach towards understanding how this critical protein works. The report illustrates stopped-flow experiments for measuring the coupled DNA binding and ATPase kinetics underlying Msh2-Msh6 mechanism of action in DNA repair.
Tags
Stopped-flow Kinetics MethodsMechanism Of ActionDNA Repair ProteinTransient Kinetic AnalysisBiological MacromoleculesActive Site ConcentrationsIntrinsic Rate ConstantsMacromolecular FunctionEnzymesPre-steady State MeasurementsReaction PathwaySteady State MeasurementsCatalytic EfficiencyCatalytic SpecificityProtein-protein InteractionsProtein-ligand InteractionsRate-limiting Conformational ChangesMillisecond TimescaleStopped-flow MethodChemical-quench Flow MethodsOptical SignalFluorescenceSubstrate BindingProduct ReleaseCatalytic TurnoverMsh2-Msh6 ProteinEukaryotic DNA Repair ProteinBase-pair MismatchesInsertion/deletion Loops In DNAMismatch Repair (MMR)Genomic Integrity
