AirID, a novel proximity biotinylation enzyme, for analysis of protein-protein interactions
- Ehime University, Japan
- University of Shizuoka, Japan
- Tokushima University, Japan
Abstract
Proximity biotinylation based on Escherichia coli BirA enzymes like BioID (BirA*) and TurboID is a key technology for identifying proteins interacting with a target protein in a cell or organism. However, there have been some improvements in the enzymes 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 like 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-IκBα biotinylated the endogenous CUL4 and RBX1 in the CRL4CRBN complex based on the streptavidin pull-down assay. LC-MS/MS analysis of cells stably expressing AirID-IκBα showed top-level biotinylation of RelA proteins. These results indicate that AirID is a novel enzyme for analysing protein–protein interactions.
Data availability
All data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided for Figures 3, 4, and 6.
Article and author information
Author details
Funding
Japan Agency for Medical Research and Development (JP19am0101077)
- Tatsuya Sawasaki
Japan Society for the Promotion of Science (JP16H06579)
- Tatsuya Sawasaki
Japan Society for the Promotion of Science (JP16H04729)
- Tatsuya Sawasaki
Japan Society for the Promotion of Science (JP19H03218)
- Tatsuya Sawasaki
Japan Society for the Promotion of Science (18KK0229)
- Hidetaka Kosako
Japan Society for the Promotion of Science (19H04966)
- Hidetaka Kosako
Takeda Science Foundation
- Tatsuya Sawasaki
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2020, Kido et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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AirID, a novel proximity biotinylation enzyme, for analysis of protein-protein interactions
eLife 9:e54983.
https://doi.org/10.7554/eLife.54983
