Dataset Information

Fishing for newly synthesized proteins with phosphonate-handles

ABSTRACT: Bioorthogonal chemistry introduces affinity-labels into biomolecules with minimal disruption to the original system and is widely applicable in a range of contexts. In proteomics, immobilized metal affinity chromatography (IMAC) enables enrichment of phosphopeptides with extreme sensitivity and selectivity. Here, we adapt and combine these superb assets in a new enrichment strategy using phosphonate-handles, which we term ‘PhosID’. In this approach, ‘click-able’ phosphonate-handles are introduced into proteins via 1,3-dipolar Huisgen-cycloaddition to azido-homo-alanine (AHA) and IMAC is then used to enrich exclusively for phosphonate-labeled peptides. In interferon-gamma (IFNγ) stimulated cells, PhosID enabled the identification of a large number of IFN responsive newly synthesized proteins (NSPs) whereby we monitored the differential synthesis of these proteins over time. Collectively, these data validate the excellent performance of PhosID with efficient analysis and quantification of hundreds of NSPs by single LC-MS/MS runs. We envision PhosID as an attractive and alternative tool for studying stimuli-sensitive proteome subsets.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Jurkat Cell, Hela Cell

SUBMITTER: wei wu

LAB HEAD: Wei Wu

PROVIDER: PXD018272 | Pride | 2020-07-28

REPOSITORIES: pride

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Dataset's files

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			Action	DRS
	20190607_F1_Ag5_Klein067_SA_FK190509_FT_sample_1_BSA.raw	Raw
	20190607_F1_Ag5_Klein067_SA_FK190509_FT_sample_2_BSA-azide.raw	Raw
	20190607_F1_Ag5_Klein067_SA_FK190509_FT_sample_3_BSA-azide_Palkyne.raw	Raw
	20190607_F1_Ag5_Klein067_SA_FK190509_FT_sample_4_BSA-azide_CuAAC.raw	Raw
	20190607_F1_Ag5_Klein067_SA_FK190509_FT_sample_5_BSA_Pdbco.raw	Raw

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Fishing for newly synthesized proteins with phosphonate-handles.

Kleinpenning Fleur F Steigenberger Barbara B Wu Wei W Heck Albert J R AJR

Nature communications 20200626 1

Bioorthogonal chemistry introduces affinity-labels into biomolecules with minimal disruption to the original system and is widely applicable in a range of contexts. In proteomics, immobilized metal affinity chromatography (IMAC) enables enrichment of phosphopeptides with extreme sensitivity and selectivity. Here, we adapt and combine these superb assets in a new enrichment strategy using phosphonate-handles, which we term PhosID. In this approach, click-able phosphonate-handles are introduced in ...[more]

PMID: 32591520

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