Asset Details
Do you wish to reserve the book?
Proximity labeling in mammalian cells with TurboID and split-TurboID
Hey, we have placed the reservation for you!
By the way, why not check out events that you can attend while you pick your title.
Oops! Something went wrong.
Looks like we were not able to place the reservation. Kindly try again later.
Are you sure you want to remove the book from the shelf?
Proximity labeling in mammalian cells with TurboID and split-TurboID
Do you wish to request the book?
Proximity labeling in mammalian cells with TurboID and split-TurboID
Please be aware that the book you have requested cannot be checked out. If you would like to checkout this book, you can reserve another copy
We have requested the book for you!
Your request is successful and it will be processed during the Library working hours. Please check the status of your request in My Requests.
Oops! Something went wrong.
Looks like we were not able to place your request. Kindly try again later.
Journal Article
Proximity labeling in mammalian cells with TurboID and split-TurboID
2020
Overview
This protocol describes the use of TurboID and split-TurboID in proximity labeling applications for mapping protein–protein interactions and subcellular proteomes in live mammalian cells. TurboID is an engineered biotin ligase that uses ATP to convert biotin into biotin–AMP, a reactive intermediate that covalently labels proximal proteins. Optimized using directed evolution, TurboID has substantially higher activity than previously described biotin ligase–related proximity labeling methods, such as BioID, enabling higher temporal resolution and broader application in vivo. Split-TurboID consists of two inactive fragments of TurboID that can be reconstituted through protein–protein interactions or organelle–organelle interactions, which can facilitate greater targeting specificity than full-length enzymes alone. Proteins biotinylated by TurboID or split-TurboID are then enriched with streptavidin beads and identified by mass spectrometry. Here, we describe fusion construct design and characterization (variable timing), proteomic sample preparation (5–7 d), mass spectrometric data acquisition (2 d), and proteomic data analysis (1 week).
This protocol describes proximity labeling approaches using TurboID and split-TurboID, which can be used for mapping protein–protein interactions and organelle proteomes in live mammalian cells with nanometer spatial resolution.
Publisher
Nature Publishing Group UK,Nature Publishing Group
