title: Evidence for sequential action of two ATPase active sites in yeast Msh2-Msh6.
abstract: Bacterial MutS homodimers contain two ATPase active sites that have non-equivalent functions in DNA mismatch repair. The homologous Msh2-Msh6 complex in eukaryotes also has intrinsic ATPase activity that is essential for mismatch repair. Here, we investigate differences in the two putative ATPase active sites by examining the properties of heterodimers containing alanine substituted for an invariant glutamic acid in the active site of either Msh2, Msh6 or both. Mutation rates in wild type versus Glu-->Ala mutant haploid yeast strains indicate that both ATPase active sites are essential for mismatch repair activity in vivo. The properties of purified heterodimers suggest that the ATPase active site in Msh6 binds ATP with higher affinity and hydrolyzes ATP faster and with higher efficiency than does the ATPase active site in Msh2. This suggests sequential action of the two ATPase active sites, in which ATP binds to Msh6 first to trigger downstream events in mismatch repair.
authors.affiliation: Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709, USA.
authors.forename: Karin
authors.initials: K
authors.lastname: Drotschmann
authors.forename: Wei
authors.initials: W
authors.lastname: Yang
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Kunkel
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D019943:Amino Acid Substitution / Q000235:genetics; D020137:Base Pair Mismatch / Q000235:genetics; D001665:Binding Sites; D001667:Binding, Competitive; D004260:DNA Repair / Q000235:genetics; D004271:DNA, Fungal / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D019281:Dimerization; D024202:Electrophoretic Mobility Shift Assay; D005656:Fungal Proteins / Q000378:metabolism*; D051718:MutS Homolog 2 Protein; D009154:Mutation / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*
chemical_list: D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C098790:MSH6 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; C489751:MSH2 protein, S cerevisiae; D051718:MutS Homolog 2 Protein