DNA Binding Properties of the Yeast Msh2-Msh6 and Mlh1-Pms1 Heterodimers

Abstract

We describe here our recent studies of the DNA binding properties of Msh2-Msh6 and Mlh1-Pms1, two protein complexes required to repair mismatches generated during DNA replication. Mismatched DNA binding by Msh2-Msh6 was probed by mutagenesis based on the crystal structure of the homologous bacterial MutS homodimer bound to DNA. The results suggest that several amino acid side chains inferred to interact with the DNA backbone near the mismatch are critical for repair activity. These contacts, which are different in Msh2 and Msh6, likely facilitate stacking and hydrogen bonding interactions between side chains in Msh6 and the mismatched base, thus stabilizing a kinked DNA conformation that permits subsequent repair steps coordinated by the Mlh1-Pms1 heterodimer. Mlh1-Pms1 also binds to DNA, but independently of a mismatch. Mlh1-Pms1 binds short DNA substrates with low affinity and with a slight preference for single-stranded DNA. It also binds longer duplex DNA molecules, but with a higher affinity indicative of cooperative binding. Indeed, imaging by atomic force microscopy reveals cooperative DNA binding and simultaneous interaction with two DNA duplexes. The novel DNA binding properties of Mlh1-Pms1 may be relevant to signal transduction during DNA mismatch repair and to recombination, meiosis and cellular responses to DNA damage.

Mismatch Repair during Homologous and Homeologous Recombination

“…The mechanism that is most consistent with the data and the Brownian nature of molecular biology appears to be the molecular switch model (Gradia et al 1997(Gradia et al , 1999Fishel 1998Acharya et al 2003;Jeong et al 2011;Cho et al 2012;Gorman et al 2012;Qiu et al 2012;Spies 2013). Most, if not all, biochemical discrepancy can be traced to differences in the experimental conditions, an issue that persists today (Hall et al 2001;Drotschmann et al 2002;Tessmer et al 2008;Sass et al 2010;Tham et al 2013).…”

“…Potential functions for EcMutL in bacterial MMR may be to stabilize the EcMutH endonuclease at the hemimethylated GATC site and/or enhance EcUvrD helicase activity during excision. Interestingly, EcMutL and ScMlh1-ScPms1 have been shown to bind ssDNA (Drotschmann et al 2002;Park et al 2010). However, EcMutL ssDNA-binding activity is nearly undetectable at physiological ionic strength and unlikely to be significant during MMR (Park et al 2010).…”

Mismatch Repair

“…The function, if any, of these conformational transitions is unknown. EcMutL and ScMlh1-ScPms1 have been shown to bind ssDNA in very low ionic strength conditions (63,64). However, this activity becomes nearly undetectable at physiological ionic strength (64).…”

Molecular Classification of Gastric Adenocarcinoma: Translating New Insights from The Cancer Genome Atlas Research Network

“…MLH1 silencing through mutation or methylation of gene & MutL homolog 1 (MLH1) is a key gene for DNA MMR, which edits errors made during DNA replication [54]. Loss of function of MLH1, mainly resulting from promoter methylation of the gene, may cause a high rate of gene mutations in the genome, leading to carcinogenesis [55].…”