title: Mispair-specific recruitment of the Mlh1-Pms1 complex identifies repair substrates of the Saccharomyces cerevisiae Msh2-Msh3 complex.
abstract: DNA mismatch repair is initiated by either the Msh2-Msh6 or the Msh2-Msh3 mispair recognition heterodimer. Here we optimized the expression and purification of Saccharomyces cerevisiae Msh2-Msh3 and performed a comparative study of Msh2-Msh3 and Msh2-Msh6 for mispair binding, sliding clamp formation, and Mlh1-Pms1 recruitment. Msh2-Msh3 formed sliding clamps and recruited Mlh1-Pms1 on +1, +2, +3, and +4 insertion/deletions and CC, AA, and possibly GG mispairs, whereas Msh2-Msh6 formed mispair-dependent sliding clamps and recruited Mlh1-Pms1 on 7 of the 8 possible base:base mispairs, the +1 insertion/deletion mispair, and to a low level on the +2 but not the +3 or +4 insertion/deletion mispairs and not on the CC mispair. The mispair specificity of sliding clamp formation and Mlh1-Pms1 recruitment but not mispair binding alone correlated best with genetic data on the mispair specificity of Msh2-Msh3- and Msh2-Msh6-dependent mismatch repair in vivo. Analysis of an Msh2-Msh6/Msh3 chimeric protein and mutant Msh2-Msh3 complexes showed that the nucleotide binding domain and communicating regions but not the mispair binding domain of Msh2-Msh3 are responsible for the extremely rapid dissociation of Msh2-Msh3 sliding clamps from DNA relative to that seen for Msh2-Msh6, and that amino acid residues predicted to stabilize Msh2-Msh3 interactions with bent, strand-separated mispair-containing DNA are more critical for the recognition of small +1 insertion/deletions than larger +4 insertion/deletions.
authors.affiliation: From the Ludwig Institute for Cancer Research.
authors.forename: Anjana
authors.initials: A
authors.lastname: Srivatsan
authors.forename: Nikki
authors.initials: N
authors.lastname: Bowen
authors.forename: Richard D
authors.initials: RD
authors.lastname: Kolodner
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D053843:DNA Mismatch Repair; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D054643:INDEL Mutation; D000070957:MutL Protein Homolog 1; D000070956:MutL Proteins; D051718:MutS Homolog 2 Protein / Q000378:metabolism*; D000074081:MutS Homolog 3 Protein; D009154:Mutation; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; C088859:MLH1 protein, S cerevisiae; C081696:MSH3 protein, S cerevisiae; D000074081:MutS Homolog 3 Protein; C088858:PMS1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C489751:MSH2 protein, S cerevisiae; D000070957:MutL Protein Homolog 1; D000070956:MutL Proteins; D051718:MutS Homolog 2 Protein
keywords: DNA Mismatch Repair; DNA Recombination; DNA Repair; DNA Replication; Mismatch Binding; Mismatch Repair Specificity; Msh6; Mutagenesis Mechanisms; Replication Fidelity