EASTBIO Defining the structural basis of Tdp1-dependent DNA repair
DNA topoisomerases are described as “magicians of the DNA world”. They allow DNA strands and double helices to “pass through each other” thereby resolving topological strain created during DNA replication and transcription. Type IB topoisomerases work their magic by forming transient covalent complexes with DNA via a tyrosine, while at the same time generating a temporary DNA single-strand break (SSB). However, if the break occurs near an existing nick, a strand gap or a mismatched base, the enzyme stalls and the DNA remains broken. Conversion of SSBs to potentially lethal double-strand breaks (DSBs) occurs during DNA replication or transcription. Cells therefore protect themselves by repairing stalled topoisomerase IB (TopIB)-DNA complexes. In eukaryotic cells, tyrosyl-DNA phosphodiesterase 1 (Tdp1) performs this important role, by hydrolysing the TopIB tyrosine-DNA bond, thus freeing the damaged DNA ends for repair. Structural snapshots of the Tdp1-dependent DNA repair pathway will provide crucial molecular-level understanding of this DNA repair mechanism, which is fundamental to maintaining human health.
Topoisomerase-based therapeutics, such as topotecan and irinotecan, stall TopIB-DNA complexes by intercalating at the TopIB-DNA junction, preventing re-ligation. Accumulation of SSBs then DSBs leads to the death of rapidly replicating cells. Tdp1 counters the action of these drugs. Therefore, blocking Tdp1’s function, with small molecule inhibitors, could improve the effectiveness of these therapies1. Understanding how Tdp1 works and how inhibitors bind, will guide the ongoing design of novel Tdp1 inhibitors.
Recently we established the structural basis of Tdp1’s ability to process 3’-DNA ends2. Our objectives now are:
1. To understand how human Tdp1 processes a physiological TopIB-DNA covalent complex.
2. To establish how Tdp1 inhibitors bind to Tdp1 and block its activity.
We use complementary structural and biochemical approaches, along with in vitro and in vivo assays developed in the Interthal lab.
The student will develop a broad range of skills in structural biology and biochemistry (in the Richardson lab) and in the molecular biology of DNA repair (in the Interthal lab).
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1. Zakharenko, A.L. et al. Novel tyrosyl-DNA phosphodiesterase 1 inhibitors enhance the therapeutic impact of topotecan on in vivo tumor models. Eur J Med Chem 161, 581-593 (2019).
2. Flett, F.J. et al. Structural basis for DNA 3'-end processing by human tyrosyl-DNA phosphodiesterase 1. Nat Commun 9, 24 (2018).
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