BioID Analysis of Actin-Binding Proteins

E Emily Joo; Michael F Olson

doi:10.1007/978-1-0716-3810-1_9

BioID Analysis of Actin-Binding Proteins

Methods Mol Biol. 2024:2794:95-104. doi: 10.1007/978-1-0716-3810-1_9.

Authors

E Emily Joo¹, Michael F Olson²

Affiliations

¹ Department of Chemistry and Biology, Toronto Metropolitan University, Toronto, ON, Canada.
² Department of Chemistry and Biology, Toronto Metropolitan University, Toronto, ON, Canada. Michael.olson@torontomu.ca.

PMID: 38630223
DOI: 10.1007/978-1-0716-3810-1_9

Abstract

Proteins often exist and function as part of higher-order complexes or networks. A challenge is to identify the universe of proximal and interacting partners for a given protein. We describe how the high-activity promiscuous biotin ligase called TurboID is fused to the actin-binding peptide LifeAct to label by biotinylation proteins that bind, or are in close proximity, to actin. The rapid enzyme kinetics of TurboID allows the profiles of actin-binding proteins to be compared under different conditions, such as acute disruption of filamentous actin structures with cytochalasin D.

Keywords: Actin; Affinity purification; Biotin ligase; Cytoskeleton; LifeAct; Proximity ligation.

MeSH terms

Actin Cytoskeleton
Actins*
Biotinylation
Microfilament Proteins*
Physics

Substances

Microfilament Proteins
Actins