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

EMBO J. 2001 Feb 1;20(3):612-8. doi: 10.1093/emboj/20.3.612.

Abstract

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.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Allosteric Regulation
  • Binding Sites
  • Crystallography, X-Ray
  • DNA, Single-Stranded / metabolism*
  • DNA-Binding Proteins / chemistry*
  • DNA-Binding Proteins / metabolism*
  • Humans
  • In Vitro Techniques
  • Models, Molecular
  • Protein Conformation
  • Protein Folding
  • Protein Structure, Tertiary
  • Replication Protein A

Substances

  • DNA, Single-Stranded
  • DNA-Binding Proteins
  • RPA1 protein, human
  • Replication Protein A