The Partner and Localizer of BRCA2 (PALB2) is a recombination mediator protein (RMP) that was discovered recently. As an RMP, it mediates DNA repair by assisting in homologous recombination, restarting replication, and DNA annealing. PALB2 has two DNA binding sites: a primary binding site, and a coil-coil domain, which interacts with BRCA1 and BRCA2, which bind to the N terminus and C terminus, respectively. Mutations within RMPs could be a source of cancer, and in this study, the mechanism by which PALB2 interacts with DNA to aid in DNA repair is examined. DNA Binding Assays and protein:DNA Stoichiometry Assays were conducted to determine the precise affinity values of the DNA to the protein. The Synergy machine measured the fluorescence anisotropy of the interaction, which allowed for the deduction of the overall binding trends of various lengths of PALB2. The use of anisotropy to measure the substrate preferences of PALB2 is unique: previous studies have used gel shift assays, but anisotropy is a more quantitative approach. The results showed that the DNA binding domain for PALB2 F1 (the first 200 amino acids of PALB2), PALB2 174 (the first 174 amino acids), and PALB2 573 (the first 573 amino acids) all have a higher affinity with double-stranded DNA (as opposed to single stranded DNA) and for longer lengths of single-stranded DNA, the stoichiometric ratio of DNA:protein binding is 1:4, which suggests that longer, single-stranded DNA can wrap around an oligomeric PALB2. These results are significant for detailing the role of PALB2 regarding its transfer of DNA from the RPA protein to the Rad51 RMP as it occurs in homologous recombination.<br/>

Mechanism of PALB2's interaction with DNA in homologous recombination

256th National Meeting (2018)

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