Specificity of the dRP/AP lyase of Ku promotes nonhomologous end joining (NHEJ) fidelity at damaged ends

Natasha Strande; Steven A Roberts; Sehyun Oh; Eric A Hendrickson; Dale A Ramsden

doi:10.1074/jbc.M111.329730

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Specificity of the dRP/AP lyase of Ku promotes nonhomologous end joining (NHEJ) fidelity at damaged ends

Journal article

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Specificity of the dRP/AP lyase of Ku promotes nonhomologous end joining (NHEJ) fidelity at damaged ends

Natasha Strande, Steven A Roberts, Sehyun Oh, Eric A Hendrickson and Dale A Ramsden

The Journal of biological chemistry, Vol.287(17), pp.13686-13693

04/20/2012

DOI: https://doi.org/10.1074/jbc.M111.329730

Handle:

https://hdl.handle.net/2376/103869

PMCID: PMC3340204

PMID: 22362780

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Abstract

Protein Structure, Tertiary

Cell Line

Oligonucleotides - chemistry

Humans

Recombinant Proteins - chemistry

Substrate Specificity

DNA - genetics

DNA Helicases - metabolism

DNA-(Apurinic or Apyrimidinic Site) Lyase - genetics

Ku Autoantigen

Recombination, Genetic

DNA End-Joining Repair

DNA Repair

DNA Damage

Proteins - chemistry

Binding Sites

Nonhomologous end joining (NHEJ) is essential for efficient repair of chromosome breaks. However, the NHEJ ligation step is often obstructed by break-associated nucleotide damage, including base loss (abasic site or 5'-dRP/AP sites). Ku, a 5'-dRP/AP lyase, can excise such damage at ends in preparation for the ligation step. We show here that this activity is greatest if the abasic site is within a short 5' overhang, when this activity is necessary and sufficient to prepare such termini for ligation. In contrast, Ku is less active near 3' strand termini, where excision would leave a ligation-blocking α,β-unsaturated aldehyde. The Ku AP lyase activity is also strongly suppressed by as little as two paired bases 5' of the abasic site. Importantly, in vitro end joining experiments show that abasic sites significantly embedded in double-stranded DNA do not block the NHEJ ligation step. Suppression of the excision activity of Ku in this context therefore is not essential for ligation and further helps NHEJ retain terminal sequence in junctions. We show that the DNA between the 5' terminus and the abasic site can also be retained in junctions formed by cellular NHEJ, indicating that these sites are at least partly resistant to other abasic site-cleaving activities as well. High levels of the 5'-dRP/AP lyase activity of Ku are thus restricted to substrates where excision of an abasic site is required for ligation, a degree of specificity that promotes more accurate joining.

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Title: Specificity of the dRP/AP lyase of Ku promotes nonhomologous end joining (NHEJ) fidelity at damaged ends
Creators: Natasha Strande - Lineberger Comprehensive Cancer Center, Department of Biochemistry and Biophysics and Curriculum in Genetics and Molecular Biology, University of North Carolina, Chapel Hill, North Carolina 27599, USA
Steven A Roberts
Sehyun Oh
Eric A Hendrickson
Dale A Ramsden
Publication Details: The Journal of biological chemistry, Vol.287(17), pp.13686-13693
Academic Unit: Molecular Biosciences, School of
Publisher: United States
Grant note: R01 CA154461 / NCI NIH HHS R01 GM088351 / NIGMS NIH HHS R01 CA084442 / NCI NIH HHS R01 CA 84442 / NCI NIH HHS
Identifiers: 99900546713301842
Language: English
Resource Type: Journal article