Two distinct DNA binding modes guide dual roles of a CRISPR-cas protein complex

Journal article (2015)

Authors

T.R. Blosser Kavli institute of nanoscience Delft, BN/Cees Dekker Lab - AS , Broad Institute of MIT and Harvard
L. Loeff Kavli institute of nanoscience Delft, BN/Chirlmin Joo Lab - AS
Edze R. Westra Wageningen University & Research, University of Exeter
Marnix Vlot Wageningen University & Research
Tim Künne Wageningen University & Research
Małgorzata Sobota Wageningen University & Research
C. Dekker Kavli institute of nanoscience Delft, BN/Cees Dekker Lab - AS
S.J.J. Brouns BN/Stan Brouns Lab - AS , Wageningen University & Research
C. Joo BN/Chirlmin Joo Lab - AS , Kavli institute of nanoscience Delft
Research Group
BN/Cees Dekker Lab (AS) (TU Delft)
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Published Date

2015

Language

English

Reuse Rights

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Faculty

Applied Sciences

Department

BN/Bionanoscience

Research Group

BN/Cees Dekker Lab

Abstract

Small RNA-guided protein complexes play an essential role in CRISPR-mediated immunity in prokaryotes. While these complexes initiate interference byflagging cognate invader DNA for destruction, recent evidence has implicated their involvement innew CRISPR memory formation, called priming, against mutated invader sequences. The mechanism by which the target recognition complex mediates these disparate responses-interference and priming-remains poorly understood. Using single-molecule FRET, we visualize how bona fide and mutated targets are differentially probed by E.coli Cascade. We observe that the recognition of bona fide targets is an ordered process that is tightly controlled forhigh fidelity. Mutated targets are recognized with low fidelity, which is featured by short-lived and PAM- and seed-independent binding by any segment of the crRNA. These dual roles of Cascade in immunity with distinct fidelities underpin CRISPR-Cas robustness, allowing for efficient degradation ofbona fide targets and priming of mutated DNA targets.