Engineering of ultraID, a compact and hyperactive enzyme for proximity-dependent biotinylation in living cells.
Commun Biol
; 5(1): 657, 2022 07 04.
Article
en En
| MEDLINE
| ID: mdl-35788163
ABSTRACT
Proximity-dependent biotinylation (PDB) combined with mass spectrometry analysis has established itself as a key technology to study protein-protein interactions in living cells. A widespread approach, BioID, uses an abortive variant of the E. coli BirA biotin protein ligase, a quite bulky enzyme with slow labeling kinetics. To improve PDB versatility and speed, various enzymes have been developed by different approaches. Here we present a small-size engineered enzyme ultraID. We show its practical use to probe the interactome of Argonaute-2 after a 10 min labeling pulse and expression at physiological levels. Moreover, using ultraID, we provide a membrane-associated interactome of coatomer, the coat protein complex of COPI vesicles. To date, ultraID is the smallest and most efficient biotin ligase available for PDB and offers the possibility of investigating interactomes at a high temporal resolution.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Ligasas de Carbono-Nitrógeno
/
Proteínas de Escherichia coli
Idioma:
En
Revista:
Commun Biol
Año:
2022
Tipo del documento:
Article
País de afiliación:
Alemania