Chapter title |
Parallel Exploration of Interaction Space by BioID and Affinity Purification Coupled to Mass Spectrometry
|
---|---|
Chapter number | 10 |
Book title |
Proteomics
|
Published in |
Methods in molecular biology, February 2017
|
DOI | 10.1007/978-1-4939-6747-6_10 |
Pubmed ID | |
Book ISBNs |
978-1-4939-6745-2, 978-1-4939-6747-6
|
Authors |
Geoffrey G. Hesketh, Ji-Young Youn, Payman Samavarchi-Tehrani, Brian Raught, Anne-Claude Gingras |
Editors |
Lucio Comai, Jonathan E. Katz, Parag Mallick |
Abstract |
Complete understanding of cellular function requires knowledge of the composition and dynamics of protein interaction networks, the importance of which spans all molecular cell biology fields. Mass spectrometry-based proteomics approaches are instrumental in this process, with affinity purification coupled to mass spectrometry (AP-MS) now widely used for defining interaction landscapes. Traditional AP-MS methods are well suited to providing information regarding the temporal aspects of soluble protein-protein interactions, but the requirement to maintain protein-protein interactions during cell lysis and AP means that both weak-affinity interactions and spatial information is lost. A more recently developed method called BioID employs the expression of bait proteins fused to a nonspecific biotin ligase, BirA*, that induces in vivo biotinylation of proximal proteins. Coupling this method to biotin affinity enrichment and mass spectrometry negates many of the solubility and interaction strength issues inherent in traditional AP-MS methods, and provides unparalleled spatial context for protein interactions. Here we describe the parallel implementation of both BioID and FLAG AP-MS allowing simultaneous exploration of both spatial and temporal aspects of protein interaction networks. |
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Mendeley readers
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