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URL: https://dx.doi.org/DOI:10.17504/protocols.io.bdrri556

Authors: Jan Pyrih, Julius Lukeš

Summary: A range of recently developed techniques that take advantage of proximity biotinylation, such as BioID, TurboID, and APEX, are particularly suitable for the studies of protein-protein interactions. As compared to classical co-immunoprecipitation, their advantage is higher reproducibility and capacity to identify stable complexes and also capture transient interactions. Briefly, the protein of interest is fused with modified biotin ligase, which promiscuously biotinylates proteins in its proximity. However, until now BioID has been used in T. brucei only in just a handful of studies. Here, we took advantage of the recently developed advanced biotin ligation-based approach named BioID2, which we have successfully adapted for T. brucei. In contrast to the classical BioID technique, BioID2 biotin ligase is smaller (26 kDa), more specific, and attached to the protein of interest by several nm-long linker arm, which improves protein folding and the biotinylation range. When applied to T. brucei, BioID2 labeling produced a highly specific output.

Associated Publications: Pyrih J, Rašková V, Škodová-Sveráková I, Pánek T, Lukeš J (2020) ZapE/Afg1 interacts with Oxa1 and its depletion causes a multifaceted phenotype. PLoS ONE 15(6): e0234918. doi: 10.1371/journal.pone.0234918

Affiliations: Institute of Parasitology, Biology Centre, Czech Academy of Sciences, České Budějovice (Budweis), Czech Republic, Institute of Parasitology, Biology Centre, Czech Academy of Sciences, České Budějovice (Budweis), Czech Republic

External URL: https://doi.org/10.1371/journal.pone.0234918

Version: 1

Publication Date: 2020

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