Identification and characterization of mitochondrial abasic (AP)-endonuclease in mammalian cells

Ranajoy Chattopadhyay, Lee Wiederhold, Bartosz Szczesny, Istvan Boldogh, Tapas K. Hazra, Tadahide Izumi, Sankar Mitra

Research output: Contribution to journalArticlepeer-review

127 Scopus citations

Abstract

Abasic (AP)-endonuclease (APE) is responsible for repair of AP sites, and single-strand DNA breaks with 3′ blocking groups that are generated either spontaneously or during repair of damaged or abnormal bases via the DNA base excision repair (BER) pathway in both nucleus and mitochondria. Mammalian cells express only one nuclear APE, 36 kDa APE1, which is essential for survival. Mammalian mitochondrial (mt) BER enzymes other than mtAPE have been characterized. In order to identify and characterize mtAPE, we purified the APE activity from beef liver mitochondria to near homogeneity, and showed that the mtAPE which has 3-fold higher specific activity relative to APE1 is derived from the latter with deletion of 33 N-terminal residues which contain the nuclear localization signal. The mtAPE-sized product could be generated by incubating 35 S-labeled APE1 with crude mitochondrial extract, but not with cytosolic or nuclear extract, suggesting that cleavage of APE1 by a specific mitochondria-associated N-terminal peptidase is a prerequisite for mitochondrial import. The low abundance of mtAPE, particularly in cultured cells might be the reason for its earlier lack of detection by western analysis.

Original languageEnglish
Pages (from-to)2067-2076
Number of pages10
JournalNucleic Acids Research
Volume34
Issue number7
DOIs
StatePublished - 2006

Bibliographical note

Funding Information:
We acknowledge Dr R. Roy’s initial studies in purification of mitochondria and characterization of APE1 antibody cross reacting proteins. We also thank Dr David Konkel for critical editing of this manuscript, and Ms. Wanda Smith for expert secretarial assistance. We are grateful to A. Kurosky and S. S. Smith of the Biomolecular Resource Facility for protein characterization. This work was supported by USPHS grants R01 CA53791, ES08457, P01 AG021830 (S.M.), CA98664 (T.I.), and NIEHS Center Grant ES06676. Funding to pay the Open Access publication charges for this article was provided by R01 CA53791.

ASJC Scopus subject areas

  • Genetics

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