Project R-12268

Substrate-Dependent Conformational Landscape of Human DNA Ligases (Research)

DNA ligases catalyze the formation of phosphodiesters on the DNA backbone and are therefore essential enzymes for DNA metabolism, such as during replication and repair. Previous research resolved the conserved modular core structure of DNA ligases and implicated that large conformational changes take place during the catalytic cycle of DNA ligases. However, it is unknown how protein flexibility regulates the enzymatic turn-over rate and their ability to engage a set of substrate structures. In this project, I will test the hypothesis that structural dynamics of DNA ligases governs substrate selectivity and enzyme turn-over rate using single-molecule FRET (smFRET) and DNA origami-tethered substrates. Fluorescently-labeled substrate DNA is used to investigate the dynamics of strand break probing by all human DNA ligases. Strategically labeled human DNA ligase III protein will subsequently be used as a model to capture the conformational landscape of DNA ligases. This project will elucidate the relationship between structural dynamics of human DNA ligases and their enzymatic activity in order to understand their functioning in different cellular contexts. In addition to fundamental insights, the gained knowledge on conformational states of DNA ligases may facilitate the understanding of disease mechanisms and the development of specific inhibitors for targeted cancer therapy.

01 November 2021 - 31 October 2023