title: Characterization of the defects in the ATP lid of E. coli MutL that cause transient hypermutability.
abstract: Mutator strains spontaneously arise in bacterial populations under stress in an attempt to increase evolutionary adaptation. Inactivation of the ubiquitous DNA mismatch repair pathway, whose normal function is to correct replication errors and hence increase replication fidelity, is often the cause of the mutator phenotype. One of the essential genes in this pathway, mutL, includes a short tandem repeat that is prone to polymerase slippage during replication. While extensive work has established that this repetitive sequence is a genuine genetic switch, the mechanism of MutL inactivation remains unclear. This short tandem repeat is translated into a LALALA motif that resides near the ATPase active site of MutL. Therefore, changes in the length of this motif are presumed to alter the ATPase activity of MutL. We have engineered variants of Escherichia coli MutL with shorter/longer LALALA motifs and characterized their ATPase and DNA binding functions. We have found that the deletion or insertion of a single LA repeat did not compromise the structural integrity of the protein, nor did it affect MutS- or DNA-binding activity. However, it severely compromised ATP binding and, consequently, engagement of the N-terminal domains; both essential activities for proper DNA mismatch repair. These results are discussed in the context of the structure of MutL.
authors.affiliation: Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, ON, Canada.
authors.forename: Monica C
authors.initials: MC
authors.lastname: Pillon
authors.forename: Michelle
authors.initials: M
authors.lastname: Dubinsky
authors.forename: Randal N
authors.initials: RN
authors.lastname: Johnston
authors.forename: Shu-Lin
authors.initials: SL
authors.lastname: Liu
authors.forename: Alba
authors.initials: A
authors.lastname: Guarné
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000737:chemistry / Q000378:metabolism*; D020816:Amino Acid Motifs; D004247:DNA / Q000378:metabolism; D053843:DNA Mismatch Repair; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D014644:Genetic Variation; D008958:Models, Molecular; D000070956:MutL Proteins; D051719:MutS DNA Mismatch-Binding Protein / Q000378:metabolism; D016296:Mutagenesis; D017510:Protein Folding; D017434:Protein Structure, Tertiary
chemical_list: D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C074770:MutL protein, E coli; D000255:Adenosine Triphosphate; D004247:DNA; D000251:Adenosine Triphosphatases; D000070956:MutL Proteins; D051719:MutS DNA Mismatch-Binding Protein; C497175:MutS protein, E coli
keywords: DNA mismatch repair; Genetic switch; K(+)–Mg(2+) tandem; MutL; Mutagenesis