Visual analysis of concerted cleavage by type IIF restriction enzyme SfiI in subsecond time region

Yuki Suzuki, Jamie L. Gilmore, Shige H. Yoshimura, Robert M. Henderson, Yuri L. Lyubchenko, Kunio Takeyasu

Research output: Contribution to journalArticlepeer-review

28 Scopus citations

Abstract

Many DNA regulatory factors require communication between distantly separated DNA sites for their activity. The type IIF restriction enzyme SfiI is often used as a model system of site communication. Here, we used fast-scanning atomic force microscopy to monitor the DNA cleavage process with SfiI and the changes in the single SfiI-DNA complex in the presence of either Mg 2+ or Ca 2+ at a scan rate of 1-2 fps. The increased time resolution allowed us to visualize the concerted cleavage of the protein at two cognate sites. The four termini generated by the cleavage were released in a multistep manner. The high temporal resolution enabled us to visualize the translocation of a DNA strand on a looped complex and intersegmental transfer of the SfiI protein in which swapping of the site is performed without protein dissociation. On the basis of our results, we propose that the SfiI tetramer can remain bound to one of the sites even after cleavage, allowing the other site on the DNA molecule to fill the empty DNA-binding cleft by combining a one-dimensional diffusion-mediated sliding and a segment transfer mechanism.

Original languageEnglish (US)
Pages (from-to)2992-2998
Number of pages7
JournalBiophysical journal
Volume101
Issue number12
DOIs
StatePublished - Dec 21 2011

ASJC Scopus subject areas

  • Biophysics

Fingerprint Dive into the research topics of 'Visual analysis of concerted cleavage by type IIF restriction enzyme SfiI in subsecond time region'. Together they form a unique fingerprint.

Cite this