Turchi, John J.Woods, Derek S.Harrington, Maureen A.Malkova, Anna L.Takagi, Yuichiro2014-07-112014-07-112013-11https://hdl.handle.net/1805/4665http://dx.doi.org/10.7912/C2/1851Indiana University-Purdue University Indianapolis (IUPUI)In mammalian cells DNA double strand breaks (DSBs) are highly variable with respect to sequence and structure all of which are recognized by the DNA- dependent protein kinase (DNA-PK), a critical component for the resolution of these breaks. Previously studies have shown that DNA-PK does not respond the same way to all DSBs but how DNA-PK senses differences in DNA substrate sequence and structure is unknown. Here we explore the enzymatic mechanism by which DNA-PK is activated by various DNA substrates. We provide evidence that recognition of DNA structural variations occur through distinct protein-protein interactions between the carboxy terminal (C-terminal) region of Ku80 and DNA-dependent protein kinase catalytic subunit (DNA-PKcs). Discrimination of terminal DNA sequences, on the other hand, occurs independently of Ku 80 C-terminal interactions and results exclusively from DNA-PKcs interactions with the DNA. We also show that sequence differences in DNA termini can drastically influence DNA repair through altered DNA-PK activation. Our results indicate that even subtle differences in DNA substrates influence DNA-PK activation and ultimately Non-homologous End Joining (NHEJ) efficiency.en-USDNA Repair,NHEJDNA-PKKu70/80Ku80 Carboxy TerminusDNA -- Research -- MethodologyDNA-protein interactionsProtein-protein interactionsDNA -- SynthesisEnzymes -- AnalysisProtein kinases -- Research -- EvaluationDNA repairDNA damageDNA-binding proteinsCellular control mechanismsNucleotide sequenceMutagenesisBiochemical geneticsThesis