CRISPR/Cas-directed programmable assembly of multi-enzyme complexes

Samuel Lim; Jiwoo Kim; Yujin Kim; Dawei Xu; Douglas S Clark

doi:10.1039/d0cc01174f

CRISPR/Cas-directed programmable assembly of multi-enzyme complexes

Chem Commun (Camb). 2020 May 5;56(36):4950-4953. doi: 10.1039/d0cc01174f.

Authors

Samuel Lim¹, Jiwoo Kim¹, Yujin Kim¹, Dawei Xu¹, Douglas S Clark²

Affiliations

¹ Department of Chemical and Biomolecular Engineering, University of California, Berkeley, CA 94720, USA. dsc@berkeley.edu.
² Department of Chemical and Biomolecular Engineering, University of California, Berkeley, CA 94720, USA. dsc@berkeley.edu and Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, 1 Cyclotron Road, Berkeley, CA 94720, USA.

PMID: 32239050
DOI: 10.1039/d0cc01174f

Abstract

We describe a versatile CRISPR/Cas-based strategy to construct multi-enzyme complexes scaffolded on a DNA template in programmable patterns. Catalytically inactive dCas9 nuclease was used in combination with SpyCatcher-SpyTag chemistry to assemble enzymes in a highly modular fashion. Five enzymes comprising the violacein biosynthesis pathway were precisely organized in nanometer proximity; a notable increase in violacein production demonstrated the benefits of scaffolding.

MeSH terms

CRISPR-Associated Protein 9 / genetics*
CRISPR-Associated Protein 9 / metabolism
CRISPR-Cas Systems / genetics*
Multienzyme Complexes / genetics*
Multienzyme Complexes / metabolism
Streptococcus pyogenes / enzymology

Substances

Multienzyme Complexes
CRISPR-Associated Protein 9