PXD031146

PXD031146 is an original dataset announced via ProteomeXchange.

Title	MicroID: A Novel Biotin Ligase Enables Rapid Proximity Ligation Proteomics
Description	Identifying protein-protein interactions is central to dissecting signaling and regulatory processes in cells. BioID is a proximity ligation method that uses a promiscuous biotin ligase to detect protein-protein interactions in cells in a highly reproducible manner. Recent advancements in proximity ligation tools have improved efficiency and timing of labeling, allowing for measurement of interactions on a cellular timescale. However, issues of size, stability, and background labeling of these constructs persist. Here we modified the structure of BioID2 to create a smaller, highly active, biotin ligase that we named MicroID. Truncation of the c-terminal of BioID2 and mutations to alleviate blockage of biotin/ATP binding at the active site of BioID2 resulted in a smaller, highly active construct with lower baseline labeling. Several additional point mutations improved the function of our modified MicroID construct compared to BioID2 and other biotin ligases, including TurboID and miniTurbo. MicroID is the smallest biotin ligase (180 AA for microID vs. 257 AA for miniTurbo and 338 AA for TurboID), yet it demonstrates only slightly less labeling activity than TurboID and outperforms miniTurboID. MicroID also had lower background labeling than TurboID. For experiments where precise temporal control of labeling is essential, we developed a MicroID mutant that has lower labeling efficiency, but significantly reduced biotin scavenging compared to BioID2. Finally, we demonstrate utility of MicroID in mass spectrometry experiments by localizing MicroID constructs to subcellular organelles and measuring protein-protein interactions.
HostingRepository	PRIDE
AnnounceDate	2022-08-12
AnnouncementXML	Submission_2022-08-11_23:56:04.398.xml
DigitalObjectIdentifier	https://dx.doi.org/10.6019/PXD031146
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Supported dataset by repository
PrimarySubmitter	James Londino
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	monohydroxylated residue; deamidated residue
Instrument	Orbitrap Fusion

Revision	Datetime	Status	ChangeLog Entry
0	2022-01-21 07:46:32	ID requested
⏵ 1	2022-08-11 23:56:05	announced

Johnson BS, Chafin L, Farkas D, Adair J, Elhance A, Farkas L, Bednash JS, Londino JD, MicroID2: A Novel Biotin Ligase Enables Rapid Proximity-Dependent Proteomics. Mol Cell Proteomics, 21(7):100256(2022) [pubmed]

submitter keyword: proximity ligation

BioID

TurboID

organelle

mass spectrometry

MicroID

James David Londino
contact affiliation	Department of Internal Medicine Division of Pulmonary, Critical Care, and Sleep Medicine Davis Heart and Lung Research Institute, 473 W. 12th Avenue, Columbus, OH. 43210
contact email	james.londino@osumc.edu
lab head
James Londino
contact affiliation	The Ohio State University
contact email	james.londino@osumc.edu
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2022/08/PXD031146

PRIDE project URI

• PRIDE
  1. PXD031146
     1. Label: PRIDE project
     2. Name: MicroID: A Novel Biotin Ligase Enables Rapid Proximity Ligation Proteomics