Abstract 1083 Visualizing the Structural Effects of Proline Variants on the BRCA1-PALB2 Binding Interface

Both Breast Cancer Gene 1 (BRCA1) and Partner and Localizer of BRCA 2 (PALB2) are critical for proper functioning of DNA repair through homologous recombination. Upon their interaction, the intrinsically disordered region (IDR) in BRCA1 folds into a coiled-coil to bind to the coiled-coil domain in PALB2. Multiple variations of unknown significance (VUS) have been documented within the coiled-coil region of each protein. This means a person carries a variation in either BRCA1 or PALB2 but there is not enough data to know the clinical significance of those variants. We'll report our findings related to the structure of three VUS and two engineered mutations located in the binding regions of BRCA1 and PALB2. Notably, all of the experimental variants are proline mutations. Because proline is known to disrupt alpha-helix formation, we hypothesize these variants will inhibit coiled-coil folding and interaction between BRCA1 and PALB2. We have also investigated the structure of a pathogenic proline variant and a benign valine variant as our positive control and negative control, respectively. We've generated constructs of each variant in vitro to determine the effect of each mutation on the structure of the protein and its ability to fold properly upon binding its respective partner. We are using circular dichroism to determine structural changes between the wild-type and mutants and in the presence of and absence of minimized binding domains for the other heterodimeric partner. Preliminary findings suggest the proline variants negatively impact the structure of BRCA1 and PALB2 to varying degrees. We will present the structural and binding insights gained from this research with a special focus given to the effect of proline substitutions on coiled-coil formation. Thank you to TCU Biology Department and the NIH for funding (R15GM135900).