Structural basis of target DNA recognition by CRISPR-Cas12k for RNA-guided DNA transposition.
Mol Cell
; 81(21): 4457-4466.e5, 2021 11 04.
Article
in En
| MEDLINE
| ID: mdl-34450043
ABSTRACT
The type V-K CRISPR-Cas system, featured by Cas12k effector with a naturally inactivated RuvC domain and associated with Tn7-like transposon for RNA-guided DNA transposition, is a promising tool for precise DNA insertion. To reveal the mechanism underlying target DNA recognition, we determined a cryoelectron microscopy (cryo-EM) structure of Cas12k from cyanobacteria Scytonema hofmanni in complex with a single guide RNA (sgRNA) and a double-stranded target DNA. Coupled with mutagenesis and in vitro DNA transposition assay, our results revealed mechanisms for the recognition of the GGTT protospacer adjacent motif (PAM) sequence and the structural elements of Cas12k critical for RNA-guided DNA transposition. These structural and mechanistic insights should aid in the development of type V-K CRISPR-transposon systems as tools for genome editing.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
DNA
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RNA
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RNA, Guide, Kinetoplastida
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Cryoelectron Microscopy
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CRISPR-Cas Systems
Language:
En
Journal:
Mol Cell
Journal subject:
BIOLOGIA MOLECULAR
Year:
2021
Document type:
Article
Affiliation country:
Publication country:
EEUU
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ESTADOS UNIDOS
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ESTADOS UNIDOS DA AMERICA
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EUA
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UNITED STATES
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UNITED STATES OF AMERICA
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US
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USA