Computational design of anti-CRISPR proteins with improved inhibition potency.
Nat Chem Biol
; 16(7): 725-730, 2020 07.
Article
in En
| MEDLINE
| ID: mdl-32284602
ABSTRACT
Anti-CRISPR (Acr) proteins are powerful tools to control CRISPR-Cas technologies. However, the available Acr repertoire is limited to naturally occurring variants. Here, we applied structure-based design on AcrIIC1, a broad-spectrum CRISPR-Cas9 inhibitor, to improve its efficacy on different targets. We first show that inserting exogenous protein domains into a selected AcrIIC1 surface site dramatically enhances inhibition of Neisseria meningitidis (Nme)Cas9. Then, applying structure-guided design to the Cas9-binding surface, we converted AcrIIC1 into AcrIIC1X, a potent inhibitor of the Staphylococcus aureus (Sau)Cas9, an orthologue widely applied for in vivo genome editing. Finally, to demonstrate the utility of AcrIIC1X for genome engineering applications, we implemented a hepatocyte-specific SauCas9 ON-switch by placing AcrIIC1X expression under regulation of microRNA-122. Our work introduces designer Acrs as important biotechnological tools and provides an innovative strategy to safeguard CRISPR technologies.
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Protein Engineering
/
MicroRNAs
/
Clustered Regularly Interspaced Short Palindromic Repeats
/
CRISPR-Cas Systems
/
Gene Editing
/
CRISPR-Associated Protein 9
Limits:
Humans
Language:
En
Journal:
Nat Chem Biol
Journal subject:
BIOLOGIA
/
QUIMICA
Year:
2020
Document type:
Article
Affiliation country: