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Updated: Jun 17, 2026

Gene Digital Circuits Based on CRISPR-Cas Systems and Anti-CRISPR Proteins
Published on: October 18, 2022
Azide-functionalized SpCas9 enables generation of site-selective and bioactive Cas9-siRNA conjugates
Danny Wilbie1, Matt Timmers1,2, Asimina Kogkalidou1
1Division of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences (UIPS), Utrecht University, Utrecht 3508 TB, The Netherlands. e.mastrobattista@uu.nl.
None:
The gene editing enzyme SpCas9 was engineered to display azide on its surface, enabling azide-alkyne click conjugation. As a proof of concept, siRNA functionalized with a reduction sensitive linker and ring-strained alkyne tetramethylthiocycloheptyne sulfoximine was conjugated to SpCas9 on four different residues with varying efficiency and retained protein activity. Conjugation to residue 539 was successful and site-selective while retaining SpCas9 and siRNA bioactivity.
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