S-glutathionylation of cysteine 99 in the APE1 protein impairs abasic endonuclease activity

Yun Jeong Kim, Daemyung Kim, Jennifer L. Illuzzi, Sarah Delaplane, Dian Su, Michel Bernier, Michael L. Gross, Millie M. Georgiadis, David M. Wilson

Research output: Contribution to journalArticle

18 Scopus citations

Abstract

Human apurinic/apyrimidinic (AP) endonuclease 1 (APE1) is a central participant in the base excision repair pathway, exhibiting AP endonuclease activity that incises the DNA backbone 5′ to an abasic site. Besides its prominent role as a DNA repair enzyme, APE1 was separately identified as a protein called redox effector factor 1, which is able to enhance the DNA binding activity of several transcription factors through a thiol-exchange-based reduction-oxidation mechanism. In the present study, we found that human APE1 is S-glutathionylated under conditions of oxidative stress both in the presence of glutathione in vitro and in cells. S-glutathionylated APE1 displayed significantly reduced AP endonuclease activity on abasic-site-containing oligonucleotide substrates, a result stemming from impaired DNA binding capacity. The combination of site-directed mutagenesis, biochemical assays, and mass spectrometric analysis identified Cys99 in human APE1 as the critical residue for the S-glutathionylation that leads to reduced AP endonuclease activity. This modification is reversible by reducing agents, which restore APE1 incision function. Our studies describe a novel posttranslational modification of APE1 that regulates the DNA repair function of the protein.

Original languageEnglish (US)
Pages (from-to)313-326
Number of pages14
JournalJournal of molecular biology
Volume414
Issue number3
DOIs
StatePublished - Dec 2 2011

Keywords

  • APEX1
  • base excision DNA repair
  • cysteine glutathionylation
  • posttranslational modification
  • redox regulation

ASJC Scopus subject areas

  • Molecular Biology

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    Kim, Y. J., Kim, D., Illuzzi, J. L., Delaplane, S., Su, D., Bernier, M., Gross, M. L., Georgiadis, M. M., & Wilson, D. M. (2011). S-glutathionylation of cysteine 99 in the APE1 protein impairs abasic endonuclease activity. Journal of molecular biology, 414(3), 313-326. https://doi.org/10.1016/j.jmb.2011.10.023