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

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

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Replication Protein A
Proliferating Cell Nuclear Antigen
DNA Repair
Repair
DNA
Proteins
Peptides
Binding Sites
DNA Glycosylases
Amino Acids
Protein Binding
Endonucleases
Guanine
Adenine
DNA-Directed DNA Polymerase
Glutathione Transferase
Titration
DNA-(Apurinic or Apyrimidinic Site) Lyase
Substitution reactions
Amino Acid Substitution

ASJC Scopus subject areas

  • Biochemistry

Cite this

Human Homolog of the MutY Repair Protein (hMYH) Physically Interacts with Proteins Involved in Long Patch DNA Base Excision Repair. / Parker, Antony; Gu, Yesong; Mahoney, William; Lee, Suk-Hee; Singh, Keshav K.; Lu, A. Lien.

In: Journal of Biological Chemistry, Vol. 276, No. 8, 23.02.2001, p. 5547-5555.

Research output: Contribution to journalArticle

Parker, Antony ; Gu, Yesong ; Mahoney, William ; Lee, Suk-Hee ; Singh, Keshav K. ; Lu, A. Lien. / Human Homolog of the MutY Repair Protein (hMYH) Physically Interacts with Proteins Involved in Long Patch DNA Base Excision Repair. In: Journal of Biological Chemistry. 2001 ; Vol. 276, No. 8. pp. 5547-5555.
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