Building A Replication Fork: Structural Synergy And Molecular Crosstalk Between Bacterial Initiators And Helicase Loaders

Abstract

The controlled initiation of DNA replication is critical to all cells. Dedicated initiator factors, which regulate this process, are members of the ATPases Associated with various cellular Activities (AAA+) superfamily, a broad grouping of evolutionarily related enzymes that remodel target macromolecules for numerous cellular transactions. In bacteria, ATP binding serves as the central event that allows the DnaA initiator to transition from a monomeric state into a large oligomeric complex that alters replication origins structure, triggers duplex melting, and facilitates replisome assembly. Using structural analyses, we show that ATP binding induces conformational changes in DnaA that permit the initiator to self-associate into an unanticipated, right-handed helical assembly. This quaternary arrangement actively wraps origin DNA into a positive supercoil about the DnaA oligomer, and frees ATPase catalytic motifs for interacting with other proteins at filament ends. Recent work on DnaC, the loading factor for the DnaB helicase, shows that this AAA+ protein is close structural homolog of DnaA that also assembles into a helical structure, and that engages the initiator in an ATP-dependent manner. Our findings provide a molecular framework for understanding how prokaryotic initiators transition between inactive monomer and functional multimer states, and implicate DnaC as an adaptor that plugs into an activated DnaA assembly to ensure the proper spatial deposition of replicative hexameric helicases onto a replication origin.