Direct observation of RuvAB-catalyzed branch migration of single Holliday junctions

Amit, Roee; Gileadi, O.; Stavans, Joel

doi:10.1073/pnas.0404332101

Cited by 48 publications

(47 citation statements)

References 36 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…3 and Movie 3, which is published as supporting information on the PNAS web site). A similar result was obtained by Stavans and colleagues (15), although their detection system monitored DNA length. Our observation supports the suggestion that pauses are caused by the dissociation of the RuvB motors or the RuvA-RuvB complex from the junction and that resumption of branch migration in either direction corresponds to the equal probabilities of reassembly of RuvB hexamers or the RuvA-RuvB complex on the two alternative sets of branches.…”

Section: Resultssupporting

confidence: 69%

See 1 more Smart Citation

Direct observation of DNA rotation during branch migration of Holliday junction DNA by Escherichia coli RuvA–RuvB protein complex

Han

Tani

Hayashi

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

The Escherichia coli RuvA-RuvB complex promotes branch migration of Holliday junction DNA, which is the central intermediate of homologous recombination. Like many DNA motor proteins, it is suggested that RuvA-RuvB promotes branch migration by driving helical rotation of the DNA. To clarify the RuvA-RuvB-mediated branch migration mechanism in more detail, we observed DNA rotation during Holliday junction branch migration by attaching a bead to one end of cruciform DNA that was fixed to a glass surface at the opposite end. Bead rotation was observed when RuvA, RuvB, and ATP were added to the solution. We measured the rotational rates of the beads caused by RuvA-RuvB-mediated branch migration at various ATP concentrations. The data provided a K m value of 65 M and a Vmax value of 1.6 revolutions per second, which corresponds to 8.3 bp per second. This real-time observation of the DNA rotation not only allows us to measure the kinetics of the RuvA-RuvB-mediated branch migration, but also opens the possibility of elucidating the branch migration mechanism in detail.

show abstract

Section: Resultssupporting

confidence: 69%

“…Recently, RuvA-RuvB-mediated Holliday junction branch migration was visualized by single-molecule analyses (13)(14)(15). The branch migration rates were measured by monitoring the height of tethered beads.…”

mentioning

confidence: 99%

Direct observation of DNA rotation during branch migration of Holliday junction DNA by Escherichia coli RuvA–RuvB protein complex

Han

Tani

Hayashi

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

show abstract

“…Previously it has been reported from different systems that RuvB proteins are involved in the holliday junction formation, branch migration and resolution of holliday junction [5][6][7]. Further studies on RuvBs confirmed their involvement in multiple cellular pathways such as cell cycle progression and RNA polymerase IIdirected transcription [8], DNA damage response, replication fork reversal [9], nonsense-mediated mRNA decay [10], insulin stimulated GLUT4 translocation [11], small nucleolar ribonucleotide protein (snoRNPs) assembly [12], cellular transformation, cancer metastasis, apoptosis, mitosis, and development [13,14].…”

mentioning

confidence: 99%

Novel RuvB nuclear ATPase is specific to intraerythrocytic mitosis during schizogony of Plasmodium falciparum

Ahmad

Singh

Afrin

et al. 2012

Molecular and Biochemical Parasitology

View full text Add to dashboard Cite

“…Although protein-mediated branch migration has been studied extensively for various enzymes (20)(21)(22) and recently at the singlemolecule level for RuvAB (23)(24)(25), the mechanics of spontaneous migration remain largely unknown. Ensemble measurements of spontaneous branch migration in homologous junctions have detected a 1,000-fold decrease in the migration rate upon the addition of Mg 2ϩ (26,27).…”

mentioning

confidence: 99%

Observing spontaneous branch migration of Holliday junctions one step at a time

McKinney

Freeman

Lilley

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Genetic recombination occurs between homologous DNA molecules via a four-way (Holliday) junction intermediate. This ancient and ubiquitous process is important for the repair of doublestranded breaks, the restart of stalled replication forks, and the creation of genetic diversity. Once formed, the four-way junction alone can undergo the stepwise exchange of base pairs known as spontaneous branch migration. Conventional ensemble assays, useful for finding average migration rates over long sequences, have been unable to examine the affect of sequence and structure on the migration process. Here, we present a single-molecule spontaneous branch migration assay with single-base pair resolution in a study of individual DNA junctions that can undergo one step of migration. Junctions exhibit markedly different dynamics of exchange between stacking conformers depending on the point of strand exchange, allowing the moment at which branch migration occurs to be detected. The free energy landscape of spontaneous branch migration is found to be highly nonuniform and governed by two types of sequence-dependent barriers, with unmediated local migration being up to 10 times more rapid than the previously deduced average rate.FRET ͉ single molecule ͉ recombination ͉ DNA structure H omologous recombination is an essential process in maintaining genomic stability, and its defects can lead to serious human diseases, including cancer. To cope with DNA damage encountered during genome duplication, a four-way (Holliday) junction is formed by joining two nearly identical DNA molecules by strand exchange (1-3). To understand the mechanisms of cellular enzymes that recognize and process the Holliday junction, it is necessary to describe the static and dynamic structural properties of their natural substrates: homologous junctions. However, because there is no way to synchronize migration, structural properties have been determined only for junctions whose branch points are fixed, either by sequence (i.e., lack of homology) (4-13), the constraints of a crystal lattice (14-17), or other mechanical constructs (18). These studies have shown that in the absence of divalent metal ions the junction adopts an extended geometry in which the arms are directed toward the corners of a square with an open central region. But upon the addition of divalent metal ions, such as magnesium, the junction folds into the stacked X-structure, a pairwise, coaxial stacking of helices to form a right-handed, antiparallel cross. There are two possible conformers that differ in which pairs of helices are stacked against each other and a single junction can sample both stacking conformers, with continual conformational exchange between them (9-12, 19).Although protein-mediated branch migration has been studied extensively for various enzymes (20)(21)(22) and recently at the singlemolecule level for RuvAB (23-25), the mechanics of spontaneous migration remain largely unknown. Ensemble measurements of spontaneous branch migration in homologous junctions have detected ...

show abstract

Direct observation of RuvAB-catalyzed branch migration of single Holliday junctions

Cited by 48 publications

References 36 publications

Direct observation of DNA rotation during branch migration of Holliday junction DNA by Escherichia coli RuvA–RuvB protein complex

Direct observation of DNA rotation during branch migration of Holliday junction DNA by Escherichia coli RuvA–RuvB protein complex

Novel RuvB nuclear ATPase is specific to intraerythrocytic mitosis during schizogony of Plasmodium falciparum

Observing spontaneous branch migration of Holliday junctions one step at a time

Contact Info

Product

Resources

About