Research Article

Structure basis for RNA-guided DNA degradation by Cascade and Cas3

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Science  07 Jun 2018:
eaat0839
DOI: 10.1126/science.aat0839

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Abstract

Type I CRISPR-Cas system features a sequential target-searching and degradation process on double-stranded DNA, by the RNA-guided Cascade complex and the nuclease-helicase fusion enzyme Cas3, respectively. Here we present a 3.7 Å resolution cryo-EM structure of the Type I-E Cascade/R-loop/Cas3 complex, poised to initiate DNA degradation. Cas3 distinguishes Cascade conformations and only captures the R-loop forming Cascade, to avoid cleaving partially complementary targets. Its nuclease domain recruits the non-target strand (NTS) DNA at a bulged region for the nicking of single-stranded DNA. An additional 4.7 Å resolution cryo-EM structure captures the post-nicking state, in which the severed NTS retracts to the helicase entrance, to be threaded for ATP-dependent processive degradation. These snapshots form the basis for understanding RNA-guided DNA degradation in Type I-E CRISPR-Cas systems.

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