An EM View of the FokI Synaptic Complex by Single Particle Analysis

Overview

Journal J Mol Biol

Publisher Elsevier

Specialties Microbiology
Molecular Biology

Date 2007 May 26

PMID 17524420

Citations 6

Authors

Eva Scheuring Vanamee

John Berriman

Aneel K Aggarwal

Affiliations

Soon will be listed here.

Abstract

FokI is a type IIS restriction endonuclease that recognizes the 5'-GGATG-3' sequence and cleaves non-specifically at 9 and 13 base-pairs away on the top and bottom strands, respectively, to produce a 5' overhang. FokI is a bipartite endonuclease with separate recognition and cleavage domains. Because of its bipartite nature, FokI has received considerable interest in generating chimeric nucleases for use in biotechnology, and recently as possible therapeutic agents in gene therapy by initiating homologous gene recombination and repair. Here we show, using single-particle electron microscopic studies, that the FokI active complex prefers a single conformation in which the subunits are arranged in a doughnut shape complex with protein-protein and possibly protein-DNA interactions stabilizing the cleavage complex. Our electron microscopy (EM) model provides new insights into the activation mechanism of FokI and how non-specific cleavage is avoided.

Citing Articles

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Modification-dependent restriction endonuclease, MspJI, flips 5-methylcytosine out of the DNA helix.

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Type II restriction endonucleases--a historical perspective and more.

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DNA looping by FokI: the impact of synapse geometry on loop topology at varied site orientations.

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Three-dimensional EM structure of an intact activator-dependent transcription initiation complex.

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