This paper reports reconstitution of 5′-nick-directed mismatch repair using purified human proteins. The reconstituted system includes MutSα or MutSβ, MutLα, RPA, EXO1, HMGB1, PCNA, RFC, polymerase δ, and ligase I. In this system, MutSβ plays a limited role in repair of base-base mismatches, but it processes insertion/deletion mispairs much more efficiently than MutSα, which efficiently corrects both types of heteroduplexes. MutLα reduces the processivity of EXO1 and terminates EXO1-catalyzed excision upon mismatch removal. In the absence of MutLα, mismatch-provoked excision by EXO1 occurs extensively. RPA and HMGB1 play similar but complementary roles in stimulating MutSα-activated, EXO1-catalyzed excision in the presence of a mismatch, but RPA has a distinct role in facilitating MutLα-mediated excision termination past mismatch. Evidence is provided that efficient repair of a single mismatch requires multiple molecules of MutSα-MutLα complex. These data suggest a model for human mismatch repair involving coordinated initiation and termination of mismatch-provoked excision.
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)