Four healthy male volunteers (age: 24 - 27; BMI: 24.2 - 25.9 kg/m2) underwent a single bout of high-intensity bicycle exercise reaching a VO2 max approximately 95% with blood samples and muscle biopsies taken immediately pre- and post-exercise. Subjects reached 95% VO2 max in approximately 9 to 12 min and, significant increases in blood glucose and lactate were observed. Quantitative phosphoproteomic analysis of muscle biopsies pre- and post-exercise was performed with tandem mass tags, phosphopeptide enrichment using titanium dioxide chromatography, sequential elution from immobilized metal ion affinity chromatography and hydrophilic interaction liquid chromatography (TiSH) (PMID: 22906719) followed by nano-ultra high pressure liquid chromatography coupled to tandem mass spectrometry (nanoUHPLC-MS/MS). The analysis included the quantification of non-phosphorylated peptides to investigate changes in protein abundance. A total of 11,903 unique phosphopeptides (8,511 phosphosites with >90% localization probability) and 4,317 protein groups were quantified in all four subjects. The phosphorylation profile in each subject was highly reproducible with an average Pearson’s correlation coefficient r = 0.72. We identified 1,322 phosphopeptides (1,004 phosphosites with >90% localization probability) that were significantly regulated with acute exercise and only 5 protein groups quantified with altered abundance (Fig. 1D; Table S3; t-test P < 0.05 and false discovery rate (FDR) < 0.01). Of the significantly regulated phosphosites, a total of 592 were annotated in the PhosphoSitePlus database (PMID: 22135298) with 412 not previously annotated, representing potentially novel phosphosphorylation sites.
[dataset license: CC0 1.0 Universal (CC0 1.0)]
Keywords: skeletal muscle ; exericse ; phosphoproteomics
Principal Investigators: (in alphabetical order) |
Benjamin Parker, The University of Sydney, N/A |
Submitting User: | ccms |
Hoffman NJ, Parker BL, Chaudhuri R, Fisher-Wellman KH, Kleinert M, Humphrey SJ, Yang P, Holliday M, Trefely S, Fazakerley DJ, Stöckli J, Burchfield JG, Jensen TE, Jothi R, Kiens B, Wojtaszewski JF, Richter EA, James DE.
Global Phosphoproteomic Analysis of Human Skeletal Muscle Reveals a Network of Exercise-Regulated Kinases and AMPK Substrates.
Cell Metab. 2015 Nov 3;22(5):922-35. Epub 2015 Oct 1.
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