biostudies-literatureRish ADU.S. Department of Health & Human Services | NIH | National Institute of General Medical SciencesNIGMS NIH HHS3549https://www.ebi.ac.uk/biostudies/studies/S-EPMC10272136biostudies-literatureUnknown14(1)The Holliday junction (HJ) is a DNA intermediate of homologous recombination, involved in many fundamental physiological processes. RuvB, an ATPase motor protein, 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, providing a comprehensive understanding of HJ branch migration. RuvB assembles into a spiral staircase, ring-like hexamer, encircling dsDNA. Four protomers of RuvB contact the DNA backbone with a translocation step size of 2 nucleotides. The variation of nucleotide-binding states in RuvB supports a sequential model for ATP hydrolysis and nucleotide recycling, which occur at separate, singular positions. RuvB's asymmetric assembly also explains the 6:4 stoichiometry between the RuvB/RuvA cNature communicationsMolecular mechanisms of Holliday junction branch migration catalyzed by an asymmetric RuvB hexamer.PMC10272136R35 GM147465P41 GM103310GM1R35GM147465-01T32 GM118291T32 GM144293Shen ZChen ZRish ADZhang NZheng QFu TMfalseMolecular mechanisms of Holliday junction branch migration catalyzed by an asymmetric RuvB hexamer.The Holliday junction (HJ) is a DNA intermediate of homologous recombination, involved in many fundamental physiological processes. RuvB, an ATPase motor protein, 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, providing a comprehensive understanding of HJ branch migration. RuvB assembles into a spiral staircase, ring-like hexamer, encircling dsDNA. Four protomers of RuvB contact the DNA backbone with a translocation step size of 2 nucleotides. The variation of nucleotide-binding states in RuvB supports a sequential model for ATP hydrolysis and nucleotide recycling, which occur at separate, singular positions. RuvB's asymmetric assembly also explains the 6:4 stoichiometry between the RuvB/RuvA c2023-01-01T00:00:00Z2023 Jun2026-05-28T12:10:41.718Z2025-02-19T00:49:58.673ZS-EPMC102721363732206910.1038/s41467-023-39250-6