Fig. 7: Configurational changes in RPA are induced by a S384D substitution within the Aurora B motif in DBD-B.

HDX changes between RPA and RPA^S384D are shown in the (a) absence or (b) presence of ssDNA (ssDNA is depicted in black). Changes in deuterium incorporation are observed in almost all DNA binding and protein-interaction domains. Data are mapped onto the structure of human RPA which is built using the structures of the OB domains from crystal structures. The regions colored yellow correspond to peptides that were not identified in the MS analysis of either or both the wild type and mutant RPA samples. The flexible linkers were modeled using AlphaFold. The position of Ser-384 is denoted in green. Data are presented as ±SDM from three independent experiments. Bio-layer interferometry analysis of RPA or RPA^S384D binding to DSS1 in the (c) absence or (d) presence of ssDNA. RPA^S384D shows reduced binding to DSS1 in the absence of DNA. When ssDNA-bound, almost complete loss of DSS1 binding to RPA is observed.