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 eukarya remains speculative. Here we report a 2.5 Angstroms structure of the ssDNA-binding domain of human replication protein A (RPA) (eukaryotic SSB), for which we previously reported a structure in complex with ssDNA. A comparison of free and bound forms of RPA revealed that ssDNA binding is associated with a major reorientation between, and significant conformational changes within, the structural modules--OB-folds--which comprise the DNA-binding domain. Two OB-folds, whose tandem orientation was stabilized by the presence of DNA, adopted multiple orientations in its absence. Within the OB-folds, extended loops implicated in DNA binding significantly changed conformation in the absence of DNA. Analysis of intermolecular contacts suggested the possibility that other RPA molecules and/or other proteins could compete with DNA for the same binding site. Using this mechanism, protein-protein interactions 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.
Bochkareva
E
E
Department of Biochemistry and Molecular Biology, The University of Oklahoma Health Sciences Center, 975 NE 10th Street, BRC-466, Oklahoma City, OK 73190, USA.
Belegu
V
V
Korolev
S
S
Bochkarev
A
A
D000494:Allosteric Regulation; D001665:Binding Sites; D018360:Crystallography, X-Ray; D004277:DNA, Single-Stranded / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D008958:Models, Molecular; D011487:Protein Conformation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D051716:Replication Protein A
D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; C497167:RPA1 protein, human; D051716:Replication Protein A