期刊
ELIFE
卷 9, 期 -, 页码 -出版社
eLIFE SCIENCES PUBL LTD
DOI: 10.7554/elife.54983
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资金
- Japan Agency for Medical Research and Development [JP19am0101077]
- Japan Society for the Promotion of Science [JP16H06579, JP16H04729, JP19H03218, 18KK0229, 19H04966]
- Takeda Science Foundation
- University of Tokushima
- Grants-in-Aid for Scientific Research [18KK0229, 19H04966] Funding Source: KAKEN
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-I kappa B alpha indicated biotinylation of MDM2 or ReIA, 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 CRL4(CRBN) complex based on the streptavidin pull-down assay. LC-MS/MS analysis of cells that were stably expressing AirID-I kappa B alpha showed top-level biotinylation of RelA proteins. These results indicate that AirID is a novel enzyme for analyzing protein-protein interactions.
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