WEHI PhD Completion Seminar hosted by Associate Professor Shabih Shakeel

Shraddha Kameshwar

PhD student – Shakeel Laboratory, Molecular Sciences division, WEHI and Deans Laboratory, St Vincent Institute

Untying the knot: A structural look at DNA untangling in a DNA repair disorder

L7W Seminar Room

Join via TEAMS

Including Q&A session

Double-strand DNA breaks (DSBs) are the most toxic forms of DNA damage, and if untreated cause chromosomal rearrangements, genome instability, and cancer. Homologous recombination is a high-fidelity pathway for DSB repair and recruits several DNA repair proteins including the topoisomerase TOP3A and two genome stability proteins, RMI1 and RMI2. Together, they form the TOP3A cleavage complex (TRR) that is responsible for untangling or dissolving interlinked DNA repair intermediates.

Being a multi-domain protein complex, several domains within TRR remain structurally and biochemically uncharacterised. Proposed dissolution mechanisms heavily focus only on the evolutionarily conserved domains within the complex. Using Hydrogen-Deuterium Exchange Mass Spectrometry (HDX-MS), I have mapped novel DNA binding sites in poorly characterised regions of the complex. Subsequent validation using various biochemical assays provide new insights on how full-length TRR engages with DNA. Cryo-electron microscopy (cryo-EM) was used to complement biochemical observations and visualise the assembly of the full-length TRR complex on a substrate DNA.

These findings add to the available knowledge on the dynamics of substrate engagement by TRR, a complex crucial for efficient and error-free repair of damaged DNA.

All welcome!