Evidence That the DNA Endonuclease ARTEMIS also Has Intrinsic 5′-Exonuclease Activity*
| Autor: | Christian Andrew Hassig, Anthony B. Pinkerton, Michael P Hedrick, Sicong Li, Doris Niewolik, Howard H.Y. Chang, Michael R. Lieber, Klaus Schwarz |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2014 |
| Předmět: |
Exonuclease
DNA End-Joining Repair DNA repair Oligonucleotides DNA and Chromosomes Transfection Biochemistry Endonuclease chemistry.chemical_compound Nucleotidases Deoxyribonuclease I Humans Point Mutation Protein–DNA interaction Molecular Biology Nuclease Chromatography biology Oligonucleotide Circular Dichroism Nuclear Proteins Cell Biology DNA Endonucleases Molecular biology DNA-Binding Proteins enzymes and coenzymes (carbohydrates) Exodeoxyribonucleases HEK293 Cells chemistry Mutagenesis biology.protein biological phenomena cell phenomena and immunity |
| Popis: | 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. |
| Databáze: | OpenAIRE |
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