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J Biol Chem. 2014 Mar 14;289(11):7825-34. doi: 10.1074/jbc.M113.544874. Epub 2014 Feb 5.

Evidence that the DNA endonuclease ARTEMIS also has intrinsic 5'-exonuclease activity.

Author information

1
From the Departments of Pathology, Biochemistry and Molecular Biology, Biological Sciences, and Molecular Microbiology and Immunology, University of Southern California Keck School of Medicine, Norris Comprehensive Cancer Center, Los Angeles, California 90089-9176.

Abstract

ARTEMIS is a member of the metallo-β-lactamase protein family. ARTEMIS has endonuclease activity at DNA hairpins and at 5'- and 3'-DNA overhangs of duplex DNA, and this endonucleolytic activity is dependent upon DNA-PKcs. There has been uncertainty about whether ARTEMIS also has 5'-exonuclease activity on single-stranded DNA and 5'-overhangs, because this 5'-exonuclease is not dependent upon DNA-PKcs. Here, we show that the 5'-exonuclease and the endonuclease activities co-purify. Second, we show that a point mutant of ARTEMIS at a putative active site residue (H115A) markedly reduces both the endonuclease activity and the 5'-exonuclease activity. Third, divalent cation effects on the 5'-exonuclease and the endonuclease parallel one another. Fourth, both the endonuclease activity and 5'-exonuclease activity of ARTEMIS can be blocked in parallel by small molecule inhibitors, which do not block unrelated nucleases. We conclude that the 5'-exonuclease is intrinsic to ARTEMIS, making it relevant to the role of ARTEMIS in nonhomologous DNA end joining.

KEYWORDS:

DNA Repair; Nonhomologous DNA End Joining; Nucleic Acid; Nucleic Acid Enzymology; Nucleic Acid Structure; Protein DNA-Interaction

PMID:
24500713
PMCID:
PMC3953294
DOI:
10.1074/jbc.M113.544874
[Indexed for MEDLINE]
Free PMC Article
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