Structural basis for recognition of 5′-phosphotyrosine adducts by Tdp2

Abstract

SummaryThe DNA repair enzyme TDP2 resolves 5′-phosphotyrosyl-DNA adducts, and is responsible for resistance to anti-cancer drugs that target covalent topoisomerase-DNA complexes. TDP2 also participates in key signaling pathways during development and tumorigenesis, and cleaves a protein-RNA linkage during picornavirus replication. The crystal structure of zebrafish TDP2 bound to DNA reveals a deep and narrow basic groove that selectively accommodates the 5′-end of single-stranded DNA in a stretched conformation. The crystal structure of the full-length C. elegans TDP2 shows that this groove can also accommodate an acidic peptide stretch in vitro, with Glu and Asp sidechains occupying the DNA backbone phosphate binding sites. This extensive molecular mimicry suggests a potential mechanism for auto-regulation and how TDP2 may interact with phosphorylated proteins in signaling. Our study provides a framework to interrogate functions of TDP2 and develop inhibitors for chemotherapeutic and antiviral applications.

Proteolytic Degradation of Topoisomerase II (Top2) Enables the Processing of Top2·DNA and Top2·RNA Covalent Complexes by Tyrosyl-DNA-Phosphodiesterase 2 (TDP2)

“…5F). These results are consistent with the crystal structures for TDP2, showing the enzyme has a narrow DNA binding pocket, optimal for accessing single-stranded DNA ends (26,27).…”

“…2 and 4) and that mild Top2 denaturation (70°C heating) (see Fig. 3) also allowed TDP2 action indicates that the active catalytic site of TDP2 can accommodate relatively large polypeptides (26,27). This result is in line with the fact that TDP2 can process some substrates with a bulky 5Ј-tyrosyl end (25) and acts as the VPg unlinkase for picornavirus replication (36).…”

“…a digoxigenin) is attached to the tyrosine, revealing the potential of TDP2 to hydrolyze tyrosyl-DNA bonds even when a peptide is attached to the tyrosine (25). Yet crystal structures show that TDP2 has a relatively tight catalytic site (26,27), suggesting Top2 might need to be proteolyzed or denatured for TDP2 to access the tyrosyl-DNA bond. Accordingly, cellular studies indicate that the 26 S proteasome is involved in the degradation of stabilized Top2cc (28,29).…”

Involvement of the host DNA-repair enzyme TDP2 in formation of the covalently closed circular DNA persistence reservoir of hepatitis B viruses

“…2B). The preference of TDP2 for 5′ phosphotyrosyl bonds is consistent with recent X-ray data (27)(28)(29), but TDP2 also is active on 3′-substrates (20,26,30). Although TDP2 likely is the only enzyme of its kind (30), complete knockout of TDP2 in chicken DT40 cells and in mice was not lethal (20), possibly reflecting the power of redundancy in DNA repair.…”

Tyrosyl-DNA Phosphodiesterase I Catalytic Mutants Reveal an Alternative Nucleophile That Can Catalyze Substrate Cleavage

“…It has also been reported that the multifunctional enzyme Tdp2 (also known as TTRAP or EAPII) is more efficient in catalyzing the resolution of 5Ј-phosphotyrosyl adducts than Tdp1 (39). However, Tdp2 is a member of the exonuclease-endonucleasephosphatase (or EEP) nuclease family that uses single Mg 2ϩ -dependent catalysis to cleave the 5-phosphotyrosine bond and functions in the nonhomologous end joining pathway (40,41). Thus, Tdp1 and Tdp2 constitute complementary DNA repair activities, with distinct reaction mechanisms (27,42,43).…”