MassIVE MSV000093453

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O-GlcNAc Containing Proteins in Normal and Idiopathic Pulmonary Fibrosis Human Fibroblasts

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Description

Isolated normal and IPF fibroblasts were homogenized in cold MilliQ water using a bullet blender. Samples were centrifuged and inhibitors were added: HALT (Thermo Fisher Scientific), Z-Pugnac (Tocris), Thiamet G (Cayman Chemicals), and benzonase (E1014, Millipore, Sigma). O-GlcNAc enzymatic labeling and protein capturing was performed as described using a Click-IT enrichment kit following the manufacturer's protocol (cat no: C33368, C33372, and C10416; Thermo Fisher Scientific). Following an alkyne agarose bead enrichment, proteins were reduced (5 mM DTT, 30 min at 37 C), alkylated (40 mM iodoacetamide, 1 h at 37C) and digested with trypsin (1:50 enzyme/protein ratio). 0.5 ug of samples were analyzed by LC-MS/MS on a Q-Exactive HF-X mass spectrometer. Data was searched using MSFragger in match between runs mode. [doi:10.25345/C5Z02ZK7J] [dataset license: CC0 1.0 Universal (CC0 1.0)]

Keywords: fibroblasts ; O-GlcNAc ; fibrosis ; glycan

Contact

Principal Investigators:
(in alphabetical order) 		Jennifer E. Kyle, Pacific Northwest National Laboratory, United States
Submitting User: alchemistmatt

Publications

Vang S, Helton ES, Guo Y, Burpee B, Rose E, Easter M, Bollenbecker S, Hirsch MJ, Matthews EL, Jones LI, Howze PH, Rajasekaran V, Denson R, Cochran P, Attah IK, Olson H, Clair G, Melkani G, Krick S, Barnes JW.
O-GlcNAc transferase regulates collagen deposition and fibrosis resolution in idiopathic pulmonary fibrosis.
Front Immunol. Epub 2024 Apr 11.

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