Journal
NATURE COMMUNICATIONS
Volume 9, Issue -, Pages -Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/s41467-017-02530-z
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Funding
- Medical Research Council Career Development Award [G0700257]
- Scottish Universities Life Sciences Alliance (SULSA) grant
- Wellcome Trust University of Edinburgh Institutional Strategic Support Fund award
- BBSRC [BJ000884]
- BBSRC [BB/J000884/1] Funding Source: UKRI
- EPSRC [EP/K039717/1] Funding Source: UKRI
- MRC [G0700257] Funding Source: UKRI
- Biotechnology and Biological Sciences Research Council [BB/J000884/1] Funding Source: researchfish
- Engineering and Physical Sciences Research Council [EP/K039717/1] Funding Source: researchfish
- Medical Research Council [G0700257] Funding Source: researchfish
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Tyrosyl-DNA phosphodiesterase (Tdp1) is a DNA 3'-end processing enzyme that repairs topoisomerase 1B-induced DNA damage. We use a new tool combining site-specific DNA-protein cross-linking with mass spectrometry to identify Tdp1 interactions with DNA. A conserved phenylalanine (F259) of Tdp1, required for efficient DNA processing in biochemical assays, cross-links to defined positions in DNA substrates. Crystal structures of Tdp1-DNA complexes capture the DNA repair machinery after 3'-end cleavage; these reveal how Tdp1 coordinates the 3'-phosphorylated product of nucleosidase activity and accommodates duplex DNA. A hydrophobic wedge splits the DNA ends, directing the scissile strand through a channel towards the active site. The F259 side-chain stacks against the -3 base pair, delimiting the junction of duplexed and melted DNA, and fixes the scissile strand in the channel. Our results explain why Tdp1 cleavage is non-processive and provide a molecular basis for DNA 3'-end processing by Tdp1.
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