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The requirement of a Ku‐IP6 (inositol hexakisphosphate) complex for efficient repair of DNA double strand breaks by non‐homologous end joining in mammals
Author(s) -
Cheung Joyce C.Y.,
Hanakahi Les A.
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.5.a292-b
Subject(s) - ku80 , ku70 , non homologous end joining , dna , mutant , dna repair , kinase , biochemistry , genome instability , chemistry , dna pkcs , homologous recombination , microbiology and biotechnology , biology , wild type , dna damage , dna binding protein , gene , transcription factor
Non‐Homologous‐End‐Joining (NHEJ) is the major pathway in mammalian cell that repairs DNA D ouble‐ S trand‐ B reaks (DSBs) and plays a critical role in maintaining genomic stability. Inositol hexakisphosphate (IP 6 ) was previously identified to stimulate NHEJ and bind to the Ku70/Ku80 heterodimer (Ku). To study the role of Ku‐IP 6 interaction in NHEJ, selected lysine residues of the putative Ku‐IP 6 binding site were mutated to alanine. Ku wild type and IP 6 ‐binding mutants were expressed using the baculovirus expression system and purified to > 90% pure. The IP 6 ‐binding mutants have 2 – 10 fold decrease in IP 6 binding compare to wild type (K d ≈ 100 nM); they also fail to complement NHEJ in vitro . Interestingly, all the Ku‐IP 6 binding mutants bind to dsDNA ends and activate DNA‐PKcs kinase activity comparable to wild type. In summary, we have identified the IP 6 binding site in Ku and this Ku‐IP 6 interaction is essential for efficient NHEJ in vitro but unrelated to its role in binding dsDNA ends or activating DNA‐PKcs kinase activity. This research was funded by the National Institutes of Health GM070639.