Processive tanslocation and DNA unwinding by individual RecBCD enzyme molecules

Piero R. Bianco, Laurence R. Brewer, Michele Corzett, Rod Balhorn, Yin Yeh, Stephen C. Kowalczykowski, Ronald J. Baskin

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Abstract

RecBCD enzyme is a processive DNA helicase 1 and nuclease 2 that participates in the repair of chromosomal DNA through homologous recombination 3,4. We have visualized directly the movement of individual RecBCD enzymes on single molecules of double-stranded DNA (dsDNA). Detection involves the optical trapping of solitary, fluorescently tagged dsDNA molecules that are attached to polystyrene beads, and their visualization by fluorescence microscopy 5,6. Both helicase translocation and DNA unwinding are monitored by the displacement of fluorescent dye from the DNA by the enzyme 7. Here we show that unwinding is both continuous and processive, occurring at a maximum rate of 972 ± 172 base pairs per second (0.30 μm s -1), with as many as 42,300 base pairs of dsDNA unwound by a single RecBCD enzyme molecule. The mean behaviour of the individual RecBCD enzyme molecules corresponds to that observed in bulk solution.

Original languageEnglish (US)
Pages (from-to)374-378
Number of pages5
JournalNature
Volume409
Issue number6818
DOIs
StatePublished - Jan 18 2001
Externally publishedYes

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Cite this

Bianco, P. R., Brewer, L. R., Corzett, M., Balhorn, R., Yeh, Y., Kowalczykowski, S. C., & Baskin, R. J. (2001). Processive tanslocation and DNA unwinding by individual RecBCD enzyme molecules. Nature, 409(6818), 374-378. https://doi.org/10.1038/35053131