title: PARP-1 enhances the mismatch-dependence of 5'-directed excision in human mismatch repair in vitro.
abstract: End-directed mismatch-provoked excision has been reconstituted in several purified systems. While 3'-directed excision displays a mismatch dependence similar to that observed in nuclear extracts (≈20-fold), the mismatch dependence of 5'-directed excision is only 3-4-fold, significantly less than that in extracts (8-10-fold). Utilizing a fractionation-based approach, we have isolated a single polypeptide that enhances mismatch dependence of reconstituted 5'-directed excision and have shown it to be identical to poly[ADP-ribose] polymerase 1 (PARP-1). Titration of reconstituted excision reactions or PARP-1-depleted HeLa nuclear extract with purified PARP-1 showed that the protein specifically enhances mismatch dependence of 5'-directed excision. Analysis of a set of PARP-1 mutants revealed that the DNA binding domain and BRCT fold contribute to the regulation of excision specificity. Involvement of the catalytic domain is restricted to its ability to poly(ADP-ribosyl)ate PARP-1 in the presence of NAD(+), likely through interference with DNA binding. Analysis of protein-protein interactions demonstrated that PARP-1 interacts with mismatch repair proteins MutSα, exonuclease 1, replication protein A (RPA), and as previously shown by others, replication factor C (RFC) and proliferating cell nuclear antigen (PCNA) as well. The BRCT fold plays an important role in the interaction of PARP-1 with the former three proteins.
authors.affiliation: Department of Biochemistry, Box 3711, Duke University Medical Center, Durham, NC 27710, United States.
authors.forename: Yiyong
authors.initials: Y
authors.lastname: Liu
authors.forename: Farid A
authors.initials: FA
authors.lastname: Kadyrov
authors.forename: Paul
authors.initials: P
authors.lastname: Modrich
mesh_terms: D000818:Animals; D020137:Base Pair Mismatch; D002460:Cell Line; D053843:DNA Mismatch Repair; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs
chemical_list: C499320:PARP1 protein, human; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases