Human Homolog of the MutY Repair Protein (hMYH) Physically Interacts with Proteins Involved in Long Patch DNA Base Excision Repair

Antony Parker, Yesong Gu, William Mahoney, Suk Hee Lee, Keshav K. Singh, A. Lien Lu

Research output: Contribution to journalArticle

179 Scopus citations

Abstract

The human MutY homolog (hMYH) is a DNA glycosylase involved in the removal of adenines or 2-hydroxyadenines misincorporated with template guanines or 7,8-dihydro-8-oxodeoxyguanines. hMYH is associated in vivo with apurinic/apyrimidinic endonuclease (APE1), proliferating cell nuclear antigen (PCNA), and replication protein A (RPA) in HeLa nuclear extracts as shown by immunoprecipitation and Western blotting. However, binding of hMYH to DNA polymerases β and δ was not detected. By using constructs containing different portions of hMYH fused to glutathione S-transferase, we have demonstrated that the APE1-binding site is at a region around amino acid residue 300, that the PCNA binding activity is located at the C terminus, and that RPA binds to the N terminus of hMYH. A peptide consisting of residues 505-527 of hMYH that contains a conserved PCNA-binding motif binds PCNA, and subsequent amino acid substitution identified Phe-518 and Phe-519 as essential residues required for PCNA binding. RPA binds to a peptide that consists of residues 6-32 of hMYH and contains a conserved RPA-binding motif. The PCNA- and RPA-binding sites of hMYH are further confirmed by peptide and antibody titration. These results suggest that hMYH repair is a long patch base excision repair pathway.

Original languageEnglish (US)
Pages (from-to)5547-5555
Number of pages9
JournalJournal of Biological Chemistry
Volume276
Issue number8
DOIs
StatePublished - Feb 23 2001

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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