Dataset Information

LFQ_TurboID-STING_Tamavidin2-REV

ABSTRACT: The STING-null RAW264.7 cells expressing TurboID-STING were treated with DMSO or 30 μg/ml DMXAA, and 500 µM biotin was added to the medium. After 1-h incubation, the cells were lysed in 6 M guanidine-HCl containing 100 mM HEPES-NaOH (pH 7.5), 10 mM TCEP, and 40 mM CAA. The lysates were dissolved by heating and sonication, and centrifuged at 20,000 g for 15 min at 4ºC. The supernatants were recovered, and proteins were purified by methanol/chloroform precipitation and solubilized by 0.1% RapiGest SF in 50 mM triethylammonium bicarbonate. After repeated sonication and vortex, the proteins were digested with trypsin at 37ºC overnight. The resultant peptide solutions were captured on MagCapture HP Tamavidin 2-REV magnetic beads in the presence of 1 mg/ml Pefabloc SC during 3 h of incubation at 4ºC. After washing with TBS five times, the biotinylated peptides were eluted with 100 µL of 1 mM biotin at 95ºC twice. The combined eluates were desalted using GL-Tip SDB, evaporated in a SpeedVac concentrato, and redissolved in 0.1% TFA and 3% ACN.

ORGANISM(S): Mus Musculus (mouse)

SUBMITTER: Hidetaka Kosako

PROVIDER: PXD018802 | JPOST Repository | Wed Jun 17 00:00:00 BST 2020

REPOSITORIES: jPOST

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Publications

BioID screening of biotinylation sites using the avidin-like protein Tamavidin 2-REV identifies global interactors of stimulator of interferon genes (STING).

Motani Kou K Kosako Hidetaka H

The Journal of biological chemistry 20200617 32

Stimulator of interferon genes (STING) mediates cytosolic DNA-induced innate immune signaling via membrane trafficking. The global identification of proteins that spatiotemporally interact with STING will provide a better understanding of its trafficking mechanisms and of STING signaling pathways. Proximity-dependent biotin identification (BioID) is a powerful technology to identify physiologically relevant protein-protein interactions in living cells. However, biotinylated peptides are rarely d ...[more]

PMID: 32554809

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Dataset Information

LFQ_TurboID-STING_Tamavidin2-REV

Publications

BioID screening of biotinylation sites using the avidin-like protein Tamavidin 2-REV identifies global interactors of stimulator of interferon genes (STING).

Similar Datasets

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