RRC ID 62848
Author Kido K, Yamanaka S, Nakano S, Motani K, Shinohara S, Nozawa A, Kosako H, Ito S, Sawasaki T.
Title AirID, a novel proximity biotinylation enzyme, for analysis of protein-protein interactions.
Journal Elife
Abstract Proximity biotinylation based on Escherichia coli BirA enzymes such as BioID (BirA*) and TurboID is a key technology for identifying proteins that interact with a target protein in a cell or organism. However, there have been some improvements in the enzymes that are used for that purpose. Here, we demonstrate a novel BirA enzyme, AirID (ancestral BirA for proximity-dependent biotin identification), which was designed de novo using an ancestral enzyme reconstruction algorithm and metagenome data. AirID-fusion proteins such as AirID-p53 or AirID-IκBα indicated biotinylation of MDM2 or RelA, respectively, in vitro and in cells, respectively. AirID-CRBN showed the pomalidomide-dependent biotinylation of IKZF1 and SALL4 in vitro. AirID-CRBN biotinylated the endogenous CUL4 and RBX1 in the CRL4CRBN complex based on the streptavidin pull-down assay. LC-MS/MS analysis of cells that were stably expressing AirID-IκBα showed top-level biotinylation of RelA proteins. These results indicate that AirID is a novel enzyme for analyzing protein-protein interactions.
Volume 9
Published 2020-5-11
DOI 10.7554/eLife.54983
PII 54983
PMID 32391793
PMC PMC7302878
MeSH Biotin / chemistry Biotin / metabolism Biotinylation Carbon-Nitrogen Ligases / chemistry* Carbon-Nitrogen Ligases / genetics Cell Survival Escherichia coli / enzymology Escherichia coli / genetics Escherichia coli Proteins / chemistry* Escherichia coli Proteins / genetics HEK293 Cells Humans Mutation Protein Engineering* Protein Interaction Mapping Protein Interaction Maps Recombinant Fusion Proteins / chemistry* Recombinant Fusion Proteins / genetics Repressor Proteins / chemistry* Repressor Proteins / genetics
IF 7.08
Human and Animal Cells 293T(RCB2202)