Journal
NATURE METHODS
Volume 15, Issue 9, Pages 715-+Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/s41592-018-0100-5
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Funding
- Shanghai Municipal Science and Technology Commission Pujiang Talents plan [15PJ1406000, SMSTC 17411951800]
- National Natural Science Foundation of China [31570767, 31670919]
- 1,000-Youth Elite Program of China
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The communication between cells and between cellular organelles is often controlled by the interaction of membrane proteins. Although many methods for the detection of protein-protein interactions (PPIs) exist, membrane PPIs remain difficult to detect. Here we developed a proximity-based tagging system, PUP-IT (pupylation-based interaction tagging), to identify membrane protein interactions. In this approach, a small protein tag, Pup, is applied to proteins that interact with a PafA-fused bait, enabling transient and weak interactions to be enriched and detected by mass spectrometry. Pup does not diffuse from the enzyme, which allows high-specificity labeling. We applied this approach to CD28, a critical costimulatory receptor for T lymphocyte activation, and identified known CD28 binding partners and multiple potential interacting proteins. In addition, we demonstrated that this method can identify the interaction between a cell surface receptor and its ligand.
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