Nucleotide and DNA-Induced Structural Transitions and the Coupling Between ATP and DNA Binding Sites in RecA

Abstract

RecA is the prototype of ATPase proteins that mediates homologous DNA recombination. RecA requires ATP to promote the binding of DNA, as both ADP- and nucleotide-free states show low DNA binding affinity. We investigated how ATP binding affects the dynamics of DNA binding Loops, and activates RecA by the interactions between β-/γ-phosphate and Walker A and Walker B motifs, as well as through the contacts made between the C-terminal and the central domains within the protein. DNA binding results in the formation of the extended, active conformation of the RecA filament, which catalyzes strand exchange. We have performed a set of molecular dynamics simulations on the active RecA, with ATP/ADP/nucleotide-free bound, to investigate the conformational transitions between the active and inactive states. Our simulations have revealed that the structural changes upon ATP binding are confined to small motifs, while the conformational changes upon DNA binding involve larger scale rearrangement of the protein, namely the rotation of monomers with respect to each other. The results suggest that ATP binding stabilizes the L1 and L2 DNA binding loops, mediated through specific residues located between the ATP and DNA binding sites that sense the presence of γ-phosphate. Furthermore, DNA binding leads to monomer rotations to form the extended conformation by affecting the interfaces of the adjacent monomers with the help of the bound nucleotide. Based on these simulations, we propose that the ATP and DNA binding enhance the binding of each other by an allosteric mechanism mediated by a series of residues between the ATP and DNA binding sites.