DNA-dependent conformational changes in the Ku heterodimer
Open accessAbstract
Ionizing radiation induces DNA double-strand breaks which are repaired by the nonhomologous end joining (NHEJ) pathway. NHEJ is initiated upon Ku binding to the DNA ends and facilitating an interaction with the DNA-dependent protein kinase catalytic subunit (DNA-PKcs). This heterotrimeric DNA-PK complex is then active as a serine/threonine protein kinase. The molecular mechanisms involved in DNA-PK activation are unknown. Considering the crucial role of Ku in this process, we therefore determined the influence of DNA binding on the structure of the Ku heterodimer. Chemical modification with NHS-biotin and mass spectrometry were used to identify sites of modification. Biotinylation of free Ku revealed several reactive lysines on Ku70 and Ku80 which were reduced or eliminated upon DNA binding. Interestingly, in the predicted C-terminal SAP domain of Ku70, biotinylation patterns were observed which suggest a structural change in this region of the protein induced by DNA binding. Limited proteolytic digests of free and DNA-bound Ku revealed a series of unique peptides, again, indicative of a change in the accessibility of the Ku70 and Ku80 C-terminal domains. A 10 kDa peptide was also identified which was preferentially generated under non-DNA-bound conditions and mapped to the C-terminus of Ku70. These results indicate a DNA-dependent movement or structural change in the C-terminal domains of Ku70 and Ku80 that may contribute to DNA-PKcs binding and activation. These results represent the first demonstration of DNA-induced changes in Ku structure and provide a framework for analysis of DNA-PKcs and the mechanism of DNA-PK activation.
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The Ku70/Ku80 heterodimer is the first protein to bind to the end of a double-strand break.
“The very first protein to bind to the end of the double-strand break is the Ku70/Ku80 heterodimer.”
From Integration of DNA damage and repair with murine double-minute 2 (Mdm2) in tumorigenesis (2012) · cites this paper as [58]
Ku70/Ku80 forms a preformed channel that slides onto DNA termini, causing a conformational change in Ku under both redox and DNA binding conditions.
“This unique protein forms a preformed channel so that it slides onto the DNA termini, causing a conformational change in Ku under both redox and DNA binding conditions [57,58].”
From Integration of DNA damage and repair with murine double-minute 2 (Mdm2) in tumorigenesis (2012) · cites this paper as [58]
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- Authors
- Lehman, J. A., Hoelz, D. J., & Turchi, J. J.
- Affiliations
- Wright State University (US)
- Corresponding author
- Lehman, Jason A.
- ORCID
- Turchi, John J.
- Venue
- Biochemistry
- Publisher
- American Chemical Society (ACS)
- Published
- 2008-03-21
- Volume
- 47
- Issue
- 15
- Pages
- 4359-4368
- ISSN
- 0006-2960 (ISSN-L)
- 1520-4995
- PMC
- PMC2432109
- OpenAlex
- W2060734146
- Cited by
- 43
- References
- 28
- Citations by year
- 2022: 1 · 2021: 4 · 2019: 1 · 2018: 3 · 2017: 4 · 2016: 3 · 2015: 2 · 2014: 4 · 2013: 4 · 2012: 4
- FWCI
- 1.64
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Locations
- Biochemistry (publishedVersion)
- PubMed (publishedVersion)
- PubMed Central (acceptedVersion)
- IUScholarWorks (Indiana University) (submittedVersion)
- Funders
- National Institutes of Health
- Indiana University Bloomington
- National Cancer Institute (CA82741, R01CA082741)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA Repair Mechanisms
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA and Nucleic Acid Chemistry
- Health Sciences › Medicine › Radiology, Nuclear Medicine and Imaging › Effects of Radiation Exposure
- Keywords
- Ku heterodimer, Ku70, Ku80, DNA-dependent protein kinase, DNA-PKcs, non-homologous end joining, DNA double-strand breaks, DNA binding, protein conformational change, biotinylation, limited proteolysis, protein kinase activation, ionizing radiation
- MeSH terms
- Ku Autoantigen, Amino Acid Sequence, DNA, DNA-Binding Proteins, Humans, Ligands, Lysine, Models, Molecular, Molecular Sequence Data, Peptides, Protein Binding, Solvents, Mass Spectrometry, Protein Structure, Tertiary, Dimerization, Biotinylation, Antigens, Nuclear
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