Two-dimensional liquid chromatography system for online top-down mass spectrometry
An online metal-free weak cation exchange-hydrophilic interaction liquid chromatography/reversed phase liquid chromatography (WCX-HILIC/RPLC) system has been developed for sensitive high-throughput top-down mass spectrometry. Analyzing posttranslational modifications (PTMs) of core histones, with focus on histone H4, tested the system. Using ~24 μg of core histones (H4, H2B, H2A and H3) purified from human fibroblasts, 41 H4 isoforms were identified, with the type and locations of PTMs unambiguously mapped for 20 of these variants. Compared to corresponding offline studies reported previously, online WCXHILIC/ RPLC platform offers significant improvement in sensitivity, with several orders of magnitude reduction in sample requirements and reduction in the overall analysis time. To the best of our knowledge, this study represents the first online two-dimensional (2D) LC-MS/MS characterization of core histone mixture at the intact protein level.
- Research Organization:
- Pacific Northwest National Lab. (PNNL), Richland, WA (United States). Environmental Molecular Sciences Lab. (EMSL)
- Sponsoring Organization:
- USDOE
- DOE Contract Number:
- AC05-76RL01830
- OSTI ID:
- 1000615
- Report Number(s):
- PNNL-SA-73468; 30441; KP1704020; TRN: US201101%%415
- Journal Information:
- Proteomics, 10(20):3610-3620, Vol. 10, Issue 20
- Country of Publication:
- United States
- Language:
- English
Similar Records
Mapping Influenza-Induced Posttranslational Modifications on Histones from CD8+ T Cells
Isolation of Histone from Sorghum Leaf Tissue for Top Down Mass Spectrometry Profiling of Potential Epigenetic Markers