Molecular Mechanisms of Holliday Junction Branch Migration Catalyzed by an Asymmetric RuvB Hexamer

The Holliday junction (HJ) is a universal DNA intermediate of homologous recombination that is involved in many fundamental physiological processes. In bacteria, RuvB, a motor protein of the AAA+ ATPase superfamily, drives branch migration of the Holliday junction with a mechanism that had yet to be elucidated. Here, we report two cryo-EM structures of RuvB in complex with DNA and nucleotides, providing a comprehecomplex, which was further purified nsive understanding of HJ branch migration. Six RuvB protomers assemble into a spiral staircase, in the shape of a ring, with DNA in the central pore. Four protomers of RuvB hexamer interact with the backbone of the DNA substrate, suggesting a pulling-and-revolving mechanism of DNA translocation with a basic step size of 2 nucleotides. Moreover, the variation of nucleotide-binding states in our RuvB hexamer supports a sequential model for ATP hydrolysis, ADP release, and ATP reloading, which occur at specific positions on the RuvB hexamer. Furthermore, the asymmetric assembly of RuvB also explains the 6:4 stoichiometry between RuvB and RuvA, which assembles into a complex to coordinate HJ migration in cells. Taken together, we provide a comprehensive framework for the mechanistic understanding of HJ branch migration facilitated by RuvB motor protein, which may be universally shared in both prokaryotic and eukaryotic organisms.