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Deep Mutational Scanning of Protein–Protein Interactions Between Partners Expressed from Their Endogenous Loci In Vivo
Journal
Methods in Molecular Biology
ISSN
10643745
Date Issued
2022-01-01
Author(s)
Dubé, Alexandre K.
Dandage, Rohan
Dibyachintan, Soham
Dionne, Ugo
Després, Philippe C.
Landry, Christian R.
Abstract
Deep mutational scanning (DMS) generates mutants of a protein of interest in a comprehensive manner. CRISPR-Cas9 technology enables large-scale genome editing with high efficiency. Using both DMS and CRISPR-Cas9 therefore allows us to investigate the effects of thousands of mutations inserted directly in the genome. Combined with protein-fragment complementation assay (PCA), which enables the quantitative measurement of protein–protein interactions (PPIs) in vivo, these methods allow for the systematic assessment of the effects of mutations on PPIs in living cells. Here, we describe a method leveraging DMS, CRISPR-Cas9, and PCA to study the effect of point mutations on PPIs mediated by protein domains in yeast.
Volume
2477
Subjects