Crystal Structure of an Octameric RuvA–Holliday Junction Complex

Abstract

Holliday junctions occur as intermediates in homologous recombination and DNA repair. In bacteria, resolution of Holliday junctions is accomplished by the RuvABC system, consisting of a junction-specific helicase complex RuvAB, which promotes branch migration, and a junction-specific endonuclease RuvC, which nicks two strands. The crystal structure of a complex between the RuvA protein of M. leprae and a synthetic four-way junction has now been determined. Rather than binding on the open surface of a RuvA tetramer as previously suggested, the DNA is sandwiched between two RuvA tetramers, which form a closed octameric shell, stabilized by a conserved tetramer-tetramer interface. Interactions between the DNA backbone and helix-hairpin-helix motifs from both tetramers suggest a mechanism for strand separation promoted by RuvA.

Crystal structure of the Holliday junction DNA in complex with a single RuvA tetramer

“…On the other hand, the conformation of the Holliday junction and the DNA-protein interface in our complex I structure appear to differ substantially from those in complex II (16). In contrast with the concave junction DNA architecture in complex I (Fig.…”

“…The complex II crystal structure also showed a distinct scheme for holding the DNA backbones on the RuvA tetramer by the HhH motifs. Actually, both of the RuvA tetramers, the upper and the lower, simultaneously contact each strand of a DNA duplex arm (16). By contrast, in complex I, the two repeated HhH motifs bridge the two phosphodiester backbones across the minor groove of the B-form DNA (Fig.…”

“…It has been proposed that the octameric complex in form II provides a stable anchor for the RuvB motor, which would drive branch migration by rotating the DNA (16). On the other hand, the RuvC dimer was found to more efficiently resolve the junction DNA in the presence of both RuvA and RuvB, implying the formation of a higher order complex, termed the RuvABC resolvasome (7,8,30).…”

“…A full understanding of the specific interactions among the RuvA, RuvB, and RuvC proteins and the Holliday junction awaits more detailed structural studies of the higher order complex at the atomic level. Structural comparison between complex I and complex II (16). Octameric complex II is superimposed onto tetrameric complex I, which contains the refined Holliday junction structure.…”

“…The crystal structure of the E. coli RuvA-Holliday junction complex in the complex I form was solved at 6-Å resolution, and an overall structural view of the complex was reported (15). More recently, the crystal structure of octameric complex II from Mycobacterium leprae has been determined at 3.0-Å resolution (16).…”