Functional Studies and Homology Modeling of Msh2-Msh3 Predict that Mispair Recognition Involves DNA Bending and Strand Separation

Abstract

The Msh2-Msh3 heterodimer recognizes various DNA mispairs, including loops of DNA ranging from 1 to 14 nucleotides and some base-base mispairs. Homology modeling of the mispair-binding domain (MBD) of Msh3 using the related Msh6 MBD revealed that mismatch recognition must be different, even though the MBD folds must be similar. Model-based point mutation alleles of Saccharomyces cerevisiae msh3 designed to disrupt mispair recognition fell into two classes. One class caused defects in repair of both small and large insertion/deletion mispairs, whereas the second class caused defects only in the repair of small insertion/deletion mispairs; mutations of the first class also caused defects in the removal of nonhomologous tails present at the ends of double-strand breaks (DSBs) during DSB repair, whereas mutations of the second class did not cause defects in the removal of nonhomologous tails during DSB repair. Thus, recognition of small insertion/deletion mispairs by Msh3 appears to require a greater degree of interactions with the DNA conformations induced by small insertion/deletion mispairs than with those induced by large insertion/deletions that are intrinsically bent and strand separated. Mapping of the two classes of mutations onto the Msh3 MBD model appears to distinguish mispair recognition regions from DNA stabilization regions.

DNA Mismatch Repair Complex MutSβ Promotes GAA·TTC Repeat Expansion in Human Cells

“…5); these small gains point to a specific role of very small loops in GAA⅐TTC repeat expansion. Furthermore, functional tests and structural modeling have indicated that small insertions (Ͻ5) occupy a unique niche in the spectrum of mismatches that MutS␤ works with (65). Insertions of this size might be more difficult for the MutS␤ protein to accommodate (65).…”

“…Furthermore, functional tests and structural modeling have indicated that small insertions (Ͻ5) occupy a unique niche in the spectrum of mismatches that MutS␤ works with (65). Insertions of this size might be more difficult for the MutS␤ protein to accommodate (65). It is possible that discrimination can occur with respect to binding short loops of differing flexibility.…”

DNA Repair Mechanisms and the Bypass of DNA Damage in Saccharomyces cerevisiae

“…The crystal structure of hMUTSb suggests two distinct modes of IDL recognition (Gupta et al 2012), and two corresponding classes of msh3 and msh2 mutants have been identified: those defective only in the repair of small IDLs and those defective in repairing large IDLs and removing nonhomologous 39 tails (Studamire et al 1999;Dowen et al 2010). Msh3 mismatch specificity can be imparted to yeast MutSa by replacing the Msh6 mismatch-binding domain (MBD) with the presumptive Msh3 MBD; the reverse domain swap, however, does not yield a functional complex (Shell et al 2007b).…”

Mispair-specific Recruitment of the Mlh1-Pms1 Complex Identifies Repair Substrates of the Saccharomyces cerevisiae Msh2-Msh3 Complex

“…Interestingly, deuterium exchange mass spectrometry has demonstrated that MutS and Saccharomyces cerevisiae Msh2-Msh6 likely form a ring on DNA lacking a mispair, with the stacking Phe contacting the DNA; the presence of a mispair stabilizes this ring through yet other contacts between MutS or Msh6 and the DNA backbone (56). More recently, molecular modeling, genetics-based structurefunction studies, and x-ray crystallography have demonstrated that although the overall structure of the mispair-bound Msh2-Msh3 is similar to Msh2-Msh6, the interactions with the mispair are different and involve bending and strand separation of the DNA by the core mispair-contacting residues and stabilization by additional contacts with the Msh3 mispair binding domain (57,58). This importance of these additional contacts to stabilize the bound and bent DNA appears to be reduced on larger insertion/deletions (57).…”