{"database":"JPOST Repository","file_versions":[{"headers":{"Content-Type":["application/json"]},"body":{"files":{"Raw":["https://storage.jpostdb.org/JPST001131/files/151218ko12-SepPak-tC18.raw","https://storage.jpostdb.org/JPST001131/files/160312ko16-RP-C18_TFA.raw","https://storage.jpostdb.org/JPST001131/files/151218ko04-SepPak-tC18.raw","https://storage.jpostdb.org/JPST001131/files/151218ko06-RP-1.raw","https://storage.jpostdb.org/JPST001131/files/151218ko13-HLB.raw","https://storage.jpostdb.org/JPST001131/files/151218ko05-HLB.raw","https://storage.jpostdb.org/JPST001131/files/160312ko10-RP-C18_TFA.raw","https://storage.jpostdb.org/JPST001131/files/151218ko16-RP-C18.raw","https://storage.jpostdb.org/JPST001131/files/151218ko07-PLS-2.raw","https://storage.jpostdb.org/JPST001131/files/160312ko18-ref_solution.raw","https://storage.jpostdb.org/JPST001131/files/160312ko09-SepPak_pH65.raw","https://storage.jpostdb.org/JPST001131/files/151218ko08-RP-C18.raw","https://storage.jpostdb.org/JPST001131/files/160312ko15-SepPak_pH65.raw","https://storage.jpostdb.org/JPST001131/files/151218ko14-RP-1.raw","https://storage.jpostdb.org/JPST001131/files/160312ko04-RP-C18_TFA.raw","https://storage.jpostdb.org/JPST001131/files/151218ko15-PLS-2.raw","https://storage.jpostdb.org/JPST001131/files/160409ko04-MIX-6plex.raw","https://storage.jpostdb.org/JPST001131/files/160312ko03-SepPak_pH65.raw","https://storage.jpostdb.org/JPST001131/files/160312ko12-ref_solution.raw","https://storage.jpostdb.org/JPST001131/files/160312ko06-ref_solution.raw"]},"type":"primary"},"statusCodeValue":200,"statusCode":"OK"}],"scores":null,"additional":{"omics_type":["Proteomics"],"submitter":["Yasushi Ishihama"],"species":["Homo Sapiens (human)"],"full_dataset_link":["https://repository.jpostdb.org/entry/JPST001131"],"submitter_affiliation":["Kyoto university"],"sample_protocol":[""],"repository":["jPOST"],"data_protocol":[""],"pubmed_abstract":["We established a workflow for highly sensitive multiplexed quantitative phosphoproteomics using a nanoscale solid-phase tandem mass tag (TMT) labeling reactor. Phosphopeptides were first enriched by titanium oxide chromatography and then labeled with isobaric TMT reagents in a StageTip packed with hydrophobic polymer-based sorbents. We found that TMT-labeled singly phosphorylated peptides tend to flow through the titanium oxide column. Therefore, TMT labeling should be performed after the enrichment step from tryptic peptides, resulting in the need for microscale reactions with small amounts of phosphopeptides. Using an optimized protocol for tens to hundreds of nanograms of phosphopeptides, we obtained a nearly 10-fold increase in sensitivity compared to the conventional solution-based TMT protocol. We demonstrate that this nanoscale phosphoproteomics protocol works for 50 μg of HeLa proteins treated with selumetinib, and we successfully quantified the selumetinib-regulated phosphorylated sites on a proteome scale. The MS raw data files have been deposited with the ProteomeXchange Consortium via the jPOST partner repository (https://jpostdb.org) with the data set identifier PXD025536."],"pubmed_title":["Nanoscale Solid-Phase Isobaric Labeling for Multiplexed Quantitative Phosphoproteomics."],"pubmed_authors":["Ogata Kosuke K, Tsai Chia-Feng CF, Ishihama Yasushi Y"],"name_synonyms":["number, scale tissue, count in organism, 10^[-9], labeling, scales, scale, n, plant peltate hair, peltate hair, presence."],"pubmed_title_synonyms":["number, count in organism, labeling, presence."],"pubmed_abstract_synonyms":["macromolecules, scale tissue, Bru, GRP1/cytohesin 1, Raw, AI461847, Ass-1, peltate hair, number, FBN, Gene, presence, CG11633, titanio, cytohesin/GRP1, Polypeptides, method, peptido, titane, reduced, sensitive, ECTOL1, GRP1, method used in an experiment, AA408052, Grp1, Gene Products, fold, Pgi, Del(8)44H, tiny, Work Flow, Gpi-1, sensitivity, WMS, Titan, Svc, oxides, peptides, Gpi-1r, Nlk, Gpi-1s, plant peltate hair, PTPSTEP, HeLa, Phi, hypoplasia, Gpi-1t, l(2)SH2 0323, labeling, COXA_DROME, Solution, OCTD, allergic reaction, ASS, Neural-specific protein-tyrosine phosphatase, column, l(2)k08110, COX, Sensitivity, peptidos, RCB0007, oxide, GPHYSD2, Chromatographies, TAG, Kunststoff, thiol methyltransferase activity, SGS, GPH, NK/GPI, small, Data Set., Gpi, polymer molecule, Oxide, Proteins, total expressed protein, Ti, stepk, MF, Amf, Peptide, ACMICD, count in organism, TPSG1, Workflows, S-adenosyl-L-methionine:thiol S-methyltransferase activity, Phosphopeptide, mOC-X, Protein, 3.1.3.48, MFS1, NK|GPI, scales, l(2)SH0323, WMS2, NK, polymer, CYH1, covalent modifier, 22Ti, Org, ORG, titanium, Col4a-1, scale, underdeveloped, Va, Step, CG11628, tend, Specificity, Gpi1-r, Gpi1-s, Striatum-enriched protein-tyrosine phosphatase, MASS, study protocol, Gpi1-t, polymers, DmelCG11628, plan specification, Protein Gene Products, Gene Proteins, CG14724, Polymer, Specificity and Sensitivity, STEP, SSKS, PRSS31, Peptid, Polypeptide, DmelCG14724, TMT, macromolecule, Proteomes, Work Flows, Bglap-rs1, Gpi1s"],"description_synonyms":["macromolecules, scale tissue, Bru, GRP1/cytohesin 1, Raw, AI461847, Ass-1, peltate hair, number, FBN, Gene, presence, CG11633, titanio, cytohesin/GRP1, Polypeptides, method, peptido, titane, reduced, sensitive, ECTOL1, GRP1, method used in an experiment, AA408052, Grp1, Gene Products, fold, Pgi, Del(8)44H, tiny, Work Flow, Gpi-1, sensitivity, WMS, Titan, Svc, oxides, peptides, Gpi-1r, Nlk, Gpi-1s, plant peltate hair, PTPSTEP, HeLa, Phi, hypoplasia, Gpi-1t, l(2)SH2 0323, labeling, COXA_DROME, Solution, OCTD, allergic reaction, ASS, reaction, Neural-specific protein-tyrosine phosphatase, column, 10^[-6], l(2)k08110, COX, n, Sensitivity, peptidos, RCB0007, oxide, GPHYSD2, Chromatographies, TAG, Kunststoff, thiol methyltransferase activity, SGS, GPH, NK/GPI, small, Data Set., Gpi, polymer molecule, Oxide, Proteins, total expressed protein, Ti, stepk, MF, Amf, Peptide, ACMICD, count in organism, TPSG1, Workflows, S-adenosyl-L-methionine:thiol S-methyltransferase activity, Phosphopeptide, mOC-X, Protein, 3.1.3.48, MFS1, NK|GPI, scales, l(2)SH0323, WMS2, NK, polymer, CYH1, covalent modifier, 22Ti, 10^[-9], Org, ORG, titanium, Col4a-1, scale, underdeveloped, Va, Step, CG11628, tend, Specificity, Gpi1-r, Gpi1-s, Striatum-enriched protein-tyrosine phosphatase, MASS, study protocol, Gpi1-t, polymers, DmelCG11628, plan specification, Protein Gene Products, Gene Proteins, CG14724, Polymer, Specificity and Sensitivity, STEP, SSKS, PRSS31, Peptid, Polypeptide, DmelCG14724, TMT, macromolecule, Proteomes, Work Flows, Bglap-rs1, Gpi1s"],"additional_accession":[]},"is_claimable":false,"name":"Nano-scale solid-phase isobaric labeling for multiplexed quantitative phosphoproteomics","description":"We established a workflow for highly sensitive multiplexed quantitative phosphoproteomics using a nano-scale solid-phase tandem mass tag (TMT) labeling reactor. Phosphopeptides were firstly enriched by titanium oxide chromatography and then labeled with isobaric TMT reagents in a StageTip packed with hydrophobic polymer-based sorbents. We found that TMT-labeled singly phosphorylated peptides tend to flow through the titanium oxide column. Therefore, TMT labeling should be performed after the enrichment step from tryptic peptides, resulting in the need for micro-scale reaction with small amounts of phosphopeptides. Using an optimized protocol for tens to hundreds of nanograms of phosphopeptides, we obtained a nearly 10-fold increase in sensitivity compared to the conventional solution-based TMT protocol. We demonstrate that this nano-scale phosphoproteomics protocol works for 50 μg of HeLa proteins treated with selumetinib, and we successfully quantified the selumetinib-regulated phosphorylated sites on a proteome scale. The MS raw data files have been deposited with the ProteomeXchange Consortium via the jPOST partner repository (http:// jpostdb.org) with the data set identifier PXD025536.","dates":{"publication":"Fri Jul 23 00:00:00 BST 2021"},"accession":"PXD025536","cross_references":{"TAXONOMY":["9606"],"pubmed":["34292731"]}}