Novel Online Three-Dimensional Separation Expands the Detectable Functional Landscape of Cellular Phosphoproteome.
Chaewon KangSunghyun HuhDowoon NamHokeun KimJiwon HongDaehee HwangSang-Won LeePublished in: Analytical chemistry (2022)
Protein phosphorylation is a prevalent post-translational modification that regulates essentially every aspect of cellular processes. Currently, liquid chromatography-tandem mass spectrometry (LC-MS/MS) with an extensive offline sample fractionation and a phosphopeptide enrichment method is a best practice for deep phosphoproteome profiling, but balancing throughput and profiling depth remains a practical challenge. We present an online three-dimensional separation method for ultradeep phosphoproteome profiling that combines an online two-dimensional liquid chromatography separation and an additional gas-phase separation. This method identified over 100,000 phosphopeptides (>60,000 phosphosites) in HeLa cells during 1.5 days of data acquisition, and the largest HeLa cell phosphoproteome significantly expanded the detectable functional landscape of cellular phosphoproteome.
Keyphrases
- liquid chromatography
- single cell
- liquid chromatography tandem mass spectrometry
- simultaneous determination
- cell cycle arrest
- mass spectrometry
- solid phase extraction
- high resolution mass spectrometry
- tandem mass spectrometry
- induced apoptosis
- primary care
- healthcare
- ms ms
- cell death
- oxidative stress
- big data
- social media
- optical coherence tomography
- cell therapy
- small molecule
- deep learning
- cell proliferation
- binding protein
- carbon dioxide