RuvAB-mediated branch migration does not involve extensive DNA opening within the RuvB hexamer.
The Escherichia coli RuvA and RuvB proteins promote the branch migration of Holliday junctions during the late stages of homologous recombination and DNA repair (reviewed in [1]). Biochemical and structural studies of the RuvAB-Holliday junction complex have shown that RuvA binds directly to the Holliday junction [2] [3] [4] [5] [6] and acts as a specificity factor that promotes the targeting of RuvB [7] [8], a hexameric ring protein that drives branch migration [9] [10] [11]. Electron microscopic visualisation of the RuvAB complex revealed that RuvA is flanked by two RuvB hexamers, which bind DNA arms that lie diametrically opposed across the junction [8]. ATP-dependent branch migration occurs as duplex DNA is pumped out through the centre of each ring. Because RuvB possesses well-conserved helicase motifs and RuvAB exhibits a 5'-3' DNA helicase activity in vitro [12], the mechanism of branch migration is thought to involve DNA opening within the RuvB ring, which provides a single strand for the unidirectional translocation of the protein along DNA. We have investigated whether the RuvB ring can translocate along duplex DNA containing a site-directed interstrand psoralen crosslink. Surprisingly, we found that the crosslink failed to inhibit branch migration. We interpret these data as evidence against a base-by-base tracking model and suggest that extensive DNA opening within the RuvB ring is not required for DNA translocation by RuvB.
George
H
H
Clare Hall Laboratories, Imperial Cancer Research Fund, South Mimms, EN6 3LD, UK.
Kuraoka
I
I
Nauman
D A
DA
Kobertz
W R
WR
Wood
R D
RD
West
S C
SC
D000255:Adenosine Triphosphate / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000502:physiology*; D001704:Biopolymers; D004247:DNA / Q000737:chemistry / Q000235:genetics; D004265:DNA Helicases; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000502:physiology*; D029968:Escherichia coli Proteins; D006868:Hydrolysis; D011995:Recombination, Genetic
D001426:Bacterial Proteins; D001704:Biopolymers; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C061260:RuvB protein, Bacteria; D000255:Adenosine Triphosphate; D004247:DNA; C071059:Holliday junction DNA helicase, E coli; D004265:DNA Helicases