PMID:9880500

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Citation

Hall, MC and Matson, SW (1999) The Escherichia coli MutL protein physically interacts with MutH and stimulates the MutH-associated endonuclease activity. J. Biol. Chem. 274:1306-12

Abstract

All possible pairwise combinations of UvrD, MutL, MutS, and MutH were tested using the yeast two-hybrid system to identify potential interactions involving mismatch repair proteins. A two-hybrid screen previously identified a physical interaction between MutL and UvrD. Although several other known interactions were not observed, a novel interaction between MutL and MutH was detected. A series of truncations from the NH2 and COOH termini of MutL demonstrated that the COOH-terminal 218 amino acids were sufficient for the two-hybrid interaction with MutH. Removal of a small number of residues from either the NH2 or COOH termini of MutH eliminated the two-hybrid interaction with MutL. Protein affinity chromatography experiments confirmed that MutL, but not MutS, physically associates with MutH. Furthermore, MutL greatly stimulated the d(GATC)-specific endonuclease activity of MutH in the absence of MutS and a mispaired base. Stimulation of the MutH-associated endonuclease activity by MutL was dependent on ATP binding but not ATP hydrolysis. Further stimulation of this reaction by MutS required the presence of a DNA mismatch and a hydrolyzable form of ATP. These results suggest that MutL activates the MutH-associated endonuclease activity through a physical interaction during methyl-directed mismatch repair in Escherichia coli.

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Keywords

Adenosine Triphosphatases; Adenosine Triphosphate/metabolism; Adenylyl Imidodiphosphate/metabolism; Bacterial Proteins/metabolism; Base Pair Mismatch; Chromatography, Affinity; DNA Repair; DNA Repair Enzymes; DNA, Bacterial/metabolism; DNA-Binding Proteins/metabolism; Endodeoxyribonucleases/metabolism; Escherichia coli; Escherichia coli Proteins; Hydrolysis

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