Structure of the major single-stranded DNA-binding domain of replication protein A suggests a dynamic mechanism for DNA binding

Although structures of single-stranded (ss)DNA-binding proteins (SSBs) have been reported with and without ssDNA, the mechanism of ssDNA binding in eu karya remains speculative. Here we report a 2.5 Angstrom structure of the s sDNA-binding domain of human replication protein A (RPA) (eukaryotic SSB), for which we previously reported a structure in complex with ssDNA, A compa rison of free and bound forms of RPA revealed that ssDNA binding is associa ted with a major reorientation between, and significant conformational chan ges within, the structural modules-OB-folds-which comprise the DNA-binding domain. Two OB-folds, whose tandem orientation was stabilized by the presen ce of DNA, adopted multiple orientations in its absence. Within the OB-fold s, extended loops implicated in DNA binding significantly changed conformat ion in the absence of DNA, Analysis of intermolecular contacts suggested th e possibility that other RPA molecules and/or other proteins could compete with DNA for the same binding site. Using this mechanism, protein-protein i nteractions can regulate, and/or be regulated by DNA binding. Combined with available biochemical data, this structure also suggested a dynamic model for the DNA-binding mechanism.