Abstract
DNA double strand breaks (DSB) are among the most lethal forms of DNA damage and, in humans, are repaired predominantly by the non-homologous end joining (NHEJ) pathway. NHEJ is initiated by the Ku70/80 het-erodimer binding free DNA termini and then recruiting the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) to form the catalytically active DNA-PK holoenzyme. The extreme C-terminus of Ku80 (Ku80CTD) has been shown to be important for in vitro stimulation of DNA-PK activity and NHEJ in vivo. To better define the mechanism by which the Ku80CTD elicits these activities, we assessed its functional and physical interactions with DNA-PKcs and Ku70/80. The results demonstrate that DNA-PKcs activity could not be complemented by addition of a Ku80CTD suggesting that the physical connection of the C-terminus to the DNA binding domain of Ku70/80 is required for DNA-PKcs activation. Analysis of protein-protein interactions revealed a low but measurable binding of the Ku80CTD for Ku70/80ΔC and for DNA-PKcs while dimer formation and the formation of higher ordered structures of the Ku80CTD was readily apparent. Ku has been shown to tether DNA termini possibly due to protein/protein interactions. Results demonstrate that the presence of the Ku80CTD stimulates this activity possibly through Ku80CTD/Ku80CTD interactions.
| Original language | English |
|---|---|
| Pages (from-to) | 36-45 |
| Number of pages | 10 |
| Journal | International Journal of Biochemistry and Molecular Biology |
| Volume | 3 |
| Issue number | 1 |
| State | Published - 2012 |
Fingerprint
Keywords
- Chemical crosslinking
- Dimerization
- DNA repair
- DNA-PK
- Ku
- NHEJ
ASJC Scopus subject areas
- Molecular Biology
- Biochemistry
Cite this
Multiple protein-protein interactions within the DNA-PK complex are mediated by the C-terminus of Ku 80. / Bennett, Sara M.; Woods, Derek S.; Pawelczak, Katherine S.; Turchi, John.
In: International Journal of Biochemistry and Molecular Biology, Vol. 3, No. 1, 2012, p. 36-45.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Multiple protein-protein interactions within the DNA-PK complex are mediated by the C-terminus of Ku 80
AU - Bennett, Sara M.
AU - Woods, Derek S.
AU - Pawelczak, Katherine S.
AU - Turchi, John
PY - 2012
Y1 - 2012
N2 - DNA double strand breaks (DSB) are among the most lethal forms of DNA damage and, in humans, are repaired predominantly by the non-homologous end joining (NHEJ) pathway. NHEJ is initiated by the Ku70/80 het-erodimer binding free DNA termini and then recruiting the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) to form the catalytically active DNA-PK holoenzyme. The extreme C-terminus of Ku80 (Ku80CTD) has been shown to be important for in vitro stimulation of DNA-PK activity and NHEJ in vivo. To better define the mechanism by which the Ku80CTD elicits these activities, we assessed its functional and physical interactions with DNA-PKcs and Ku70/80. The results demonstrate that DNA-PKcs activity could not be complemented by addition of a Ku80CTD suggesting that the physical connection of the C-terminus to the DNA binding domain of Ku70/80 is required for DNA-PKcs activation. Analysis of protein-protein interactions revealed a low but measurable binding of the Ku80CTD for Ku70/80ΔC and for DNA-PKcs while dimer formation and the formation of higher ordered structures of the Ku80CTD was readily apparent. Ku has been shown to tether DNA termini possibly due to protein/protein interactions. Results demonstrate that the presence of the Ku80CTD stimulates this activity possibly through Ku80CTD/Ku80CTD interactions.
AB - DNA double strand breaks (DSB) are among the most lethal forms of DNA damage and, in humans, are repaired predominantly by the non-homologous end joining (NHEJ) pathway. NHEJ is initiated by the Ku70/80 het-erodimer binding free DNA termini and then recruiting the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) to form the catalytically active DNA-PK holoenzyme. The extreme C-terminus of Ku80 (Ku80CTD) has been shown to be important for in vitro stimulation of DNA-PK activity and NHEJ in vivo. To better define the mechanism by which the Ku80CTD elicits these activities, we assessed its functional and physical interactions with DNA-PKcs and Ku70/80. The results demonstrate that DNA-PKcs activity could not be complemented by addition of a Ku80CTD suggesting that the physical connection of the C-terminus to the DNA binding domain of Ku70/80 is required for DNA-PKcs activation. Analysis of protein-protein interactions revealed a low but measurable binding of the Ku80CTD for Ku70/80ΔC and for DNA-PKcs while dimer formation and the formation of higher ordered structures of the Ku80CTD was readily apparent. Ku has been shown to tether DNA termini possibly due to protein/protein interactions. Results demonstrate that the presence of the Ku80CTD stimulates this activity possibly through Ku80CTD/Ku80CTD interactions.
KW - Chemical crosslinking
KW - Dimerization
KW - DNA repair
KW - DNA-PK
KW - Ku
KW - NHEJ
UR - http://www.scopus.com/inward/record.url?scp=84859707835&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84859707835&partnerID=8YFLogxK
M3 - Article
C2 - 22509479
AN - SCOPUS:84859707835
VL - 3
SP - 36
EP - 45
JO - International Journal of Biochemistry and Molecular Biology
JF - International Journal of Biochemistry and Molecular Biology
SN - 2152-4114
IS - 1
ER -