Mechanism of 5′ Topoisomerase II DNA adduct repair by mammalian Tyrosyl DNA phosphodiesterase 2 (Tdp2)

The Topoisomerase II (topo II) DNA incision and ligation cycle can be poisoned (e.g following treatment with cancer chemotherapeutics) to generate cytotoxic DNA double strand breaks (DSBs) with topo II covalently conjugated to DNA. Tyrosyl-DNA phosphodiesterase 2 (Tdp2) protects genomic integrity by reversing 5′-phosphotyrosyl (5′-Y) linked topo II-DNA adducts. Here, X-ray structures of mouse Tdp2-DNA complexes reveal that a Tdp2 β-2-helix-β DNA damage binding “grasp”, helical “cap”, and DNA lesion binding elements fuse to form an elongated protein-DNA conjugate substrate interaction groove. The Tdp2 DNA binding surface is highly tailored for engagement of 5′-adducted ssDNA ends, and restricts non-specific endonucleolytic or exonucleolytic processing. Structural, mutational and functional analyses support a single-metal ion catalytic mechanism for the endonuclease–exonuclease–phosphatase (EEP) nuclease superfamily, and establish a molecular framework for targeted small molecule blockade of Tdp2-mediated resistance to anti-cancer topoisomerase drugs.