SARPBAXZ · Structural analysis of Rad51 paralogues involved in recombinational DNA repair

Homologous recombination (HR) is a major mechanism for repairing DNA double-strand breaks (DSBs). A DSB converts one normal chromosome into two pathological chromosomes, making it the severest form of DNA damage. Accordingly, defects in HR lead to genome instability, a potent driver of tumorigenesis. Although Rad51 is the central enzyme involved in HR, several auxiliary factors promote HR by stimulating Rad51. Rad51 paralogs comprise one family of evolutionarily conserved auxiliary factor. Unlike other auxiliary factors, Rad51 paralogs are notorious for their biochemical intractability, a trait that has greatly hindered progress in understanding their function, which in turn has precluded a more complete understanding of HR as a whole. This proposal aims to uncover the molecular mechanisms underlying Rad51 potentiation by Rad51 paralogs. To achieve this, we will employ an interdisciplinary approach combining structural biology, biophysics, biochemistry, and genetics. This proposal is particularly well-placed to deliver novel insights because the historical obstacle in the analysis of Rad51 paralogs—the native purification of functional protein of sufficient yield and quantity—has already been overcome by the Researcher. Furthermore, the Supervisor has extensive expertise in cryo-electron microscopy, which is now a viable approach to visualise the finer structural features of proteins that have been recalcitrant to classical techniques such as X-ray crystallography. Thus, by uniting the experiences of the Researcher and Supervisor, this fellowship has the potential to answer long-standing and significant questions in the field of HR. In addition to maintaining genome stability, HR plays critical roles in gametogenesis, chromosomal biology, and evolution. If awarded, this proposal therefore has the potential to impact several disciplines within the life sciences, and this is especially true given the interdisciplinary nature of the proposed research.