Structure-function analysis of CUL2LRR1 ubiquitin ligase

DNA replication is precisely regulated with unrepaired errors leading to severe pathologies including: genetic disease, cancer and premature ageing. It is essential for genomic stability that during replication active replication machinery is protected and maintained on DNA, while the machinery that has terminated is efficiently unloaded. We have shown that this unloading is driven by ubiquitylation of terminated helicase by CUL2LRR1 ubiquitin ligase. This project will use structural biology to determine, in atomic detail, how CUL2LRR1 specifically targets just the terminated replicative machinery. In collaboration with the Research Complex at Harwell we will determine X-ray and in solution structures of physiologically relevant complexes containing CUL2LRR1. We will also use cross-linking mass spectrometry to determine how CUL2LRR1 interacts with terminated replication machinery. Finally, we will validate our structures and observations in Xenopus laevis egg extract and human cell lines. This project will allow apply cutting edge structural biology techniques to the process of replication termination to better understand how specificity of interactions between protein complexes protects stability of our genomes.