title: The Saccharomyces cerevisiae Msh2 and Msh6 proteins form a complex that specifically binds to duplex oligonucleotides containing mismatched DNA base pairs.
abstract: The yeast Saccharomyces cerevisiae encodes six proteins, Msh1p to Msh6p, that show strong amino acid sequence similarity to MutS, a central component of the bacterial mutHLS mismatch repair system. Recent studies with humans and S. cerevisiae suggest that in eukaryotes, specific MutS homolog complexes that display unique DNA mismatch specificities exist. In this study, the S. cerevisiae 109-kDa Msh2 and 140-kDa Msh6 proteins were cooverexpressed in S. cerevisiae and shown to interact in an immunoprecipitation assay and by conventional chromatography. Deletion analysis of MSH2 indicated that the carboxy-terminal 114 amino acids of Msh2p are important for Msh6p interaction. Purified Msh2p-Msh6p selectively bound to duplex oligonucleotide substrates containing a G/T mismatch and a +1 insertion mismatch but did not show specific binding to +2 and +4 insertion mismatches. The mismatch binding specificity of the Msh2p-Msh6p complex, as measured by on-rate and off-rate binding studies, was abolished by ATP. Interestingly, palindromic substrates that are poorly repaired in vivo were specifically recognized by Msh2p-Msh6p; however, the binding of Msh2p-Msh6p to these substrates was not modulated by ATP. Taken together, these studies suggest that the repair of a base pair mismatch by the Msh2p-Msh6p complex is dependent on the ability of the Msh2p-Msh6p-DNA mismatch complex to use ATP hydrolysis to activate downstream events in mismatch repair.
authors.affiliation: Section of Genetics and Development, Cornell University, Ithaca, New York 14853-2703, USA. eea3@cornell.edu
authors.forename: E
authors.initials: E
authors.lastname: Alani
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism; D001482:Base Composition; D001483:Base Sequence; D001667:Binding, Competitive; D017931:DNA Primers; D004260:DNA Repair; D004271:DNA, Fungal / Q000737:chemistry* / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D005656:Fungal Proteins / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D051718:MutS Homolog 2 Protein; D016254:Mutagenesis, Insertional; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D013379:Substrate Specificity
chemical_list: D017931:DNA Primers; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C098790:MSH6 protein, S cerevisiae; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D000251:Adenosine Triphosphatases; C489751:MSH2 protein, S cerevisiae; D051718:MutS Homolog 2 Protein