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Related Experiment Video

Updated: Mar 9, 2026

In Vivo Proximity Biotinylation for Protein Interaction Studies in Paramecium tetraurelia

In Vivo Proximity Biotinylation for Protein Interaction Studies in Paramecium tetraurelia

Published on: September 12, 2025

Split-BioID: a proximity biotinylation assay for dimerization-dependent protein interactions.

Sofie De Munter¹, Janina Görnemann¹, Rita Derua^2,3

¹Laboratory of Biosignaling & Therapeutics, KU Leuven Department of Cellular and Molecular Medicine, University of Leuven, Belgium.

|December 30, 2016

Summary

This summary is machine-generated.

Split-BioID reconstitutes biotin ligase activity using inactive enzyme halves, enabling protein interaction screening. This method identifies substrates and transient interactors of protein dimers.

Keywords:

biotinylation phosphatase-substrate mapping protein-ligand screening proximity labeling reporter-fragment complementation

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Last Updated: Mar 9, 2026

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Area of Science:

Biochemistry
Molecular Biology
Proteomics

Background:

The biotin identification (BioID) protocol utilizes a modified biotin ligase (BirA*) fused to a protein of interest to label nearby proteins within living cells.
This technique is crucial for identifying protein-protein interactions and understanding cellular mechanisms.

Discussion:

This study introduces split-BioID, a novel BirA* fragment complementation approach.
Inactive halves of BirA* are fused to protein phosphatase 1 (PP1) subunits, reconstituting enzymatic activity upon PP1 heterodimerization.
This strategy allows for the specific biotinylation of proteins proximal to the reconstituted BirA* enzyme.

Key Insights:

Split-BioID successfully reconstitutes a functional BirA* enzyme through fragment complementation mediated by protein dimerization.
The split-BioID system can be effectively employed to screen for substrates and interactors of protein phosphatase 1 holoenzymes.
This method offers a versatile tool for identifying both stable and transient protein interactors, particularly within dimeric protein complexes.

Outlook:

Split-BioID provides a powerful new avenue for dissecting protein-protein interaction networks.
Further applications may include studying dynamic protein complexes and signaling pathways.
This technique holds promise for advancing our understanding of complex cellular machinery.