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Updated: Mar 30, 2026

Cell Surface Receptor Identification Using Genome-Scale CRISPR/Cas9 Genetic Screens
Published on: June 6, 2020
Gγ recruitment systems specifically select PPI and affinity-enhanced candidate proteins that interact with membrane
Misato Kaishima1, Jun Ishii2, Nobuo Fukuda3
1Department of Chemical Science and Engineering, Graduate School of Engineering, Kobe University, 1-1 Rokkodai, Nada, Kobe, Japan.
Abstract:
Protein-protein interactions (PPIs) are crucial for the vast majority of biological processes. We previously constructed a Gγ recruitment system to screen PPI candidate proteins and desirable affinity-altered (affinity-enhanced and affinity-attenuated) protein variants. The methods utilized a target protein fused to a mutated G-protein γ subunit (Gγcyto) lacking the ability to localize to the inner leaflet of the plasma membrane. However, the previous systems were adapted to use only soluble cytosolic proteins as targets. Recently, membrane proteins have been found to form the principal nodes of signaling involved in diseases and have attracted a great deal of interest as primary drug targets. Here, we describe new protocols for the Gγ recruitment systems that are specifically designed to use membrane proteins as targets to overcome previous limitations. These systems represent an attractive approach to exploring novel interacting candidates and affinity-altered protein variants and their interactions with proteins on the inner side of the plasma membrane, with high specificity and selectivity.
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