@article {doi:10.1002/elps.201900452, title = {Bottom-up proteomics of envelope proteins extracted from spinach chloroplast via high-organic-content capillary electrophoresis-mass spectrometry}, journal = {ELECTROPHORESIS}, volume = {41}, number = {n/a}, year = {2020}, month = {01/2020}, pages = {370-378}, type = {Research}, chapter = {370}, abstract = {
A high-organic-content capillary electrophoresis-tandem mass spectrometry (HOCE-MS/MS) method was developed for the proteomic analysis of envelope proteins extracted from spinach leaves. Separation was performed in a 1-meter long hydroxypropyl cellulose (HPC)-coated capillary, using 8\% (V/V) formic acid (FA) in 70\% (V/V) methanol and 22\% water as the background electrolyte (BGE). A flow-through microvial interface was used to couple the CE system with an Orbitrap Fusion Lumos mass spectrometer, and field-amplified sample stacking was used to improve the concentration sensitivity. Using this optimized method, 3579 peptides and 1141 proteins were identified using the Proteome Discoverer software with a 1\% false discovery rate at the protein level. Relative to conventional aqueous CE, HOCE-MS did a better job of discovering hydrophobic peptides and provided more peptide and protein identifications. Relative to NanoLC-MS, it achieved comparable peptide and protein identification performance and detected peptides not identified by LC-MS: of the full set of peptides identified using the two techniques, 19\% were identified only using HOCE-MS. It also outperformed NanoLC-MS with respect to the detection of low molecular weight peptides. This article is protected by copyright. All rights reserved
}, keywords = {high-organic-content capillary electrophoresis, hydrophobic peptides, proteomics of envelope proteins, spinach chloroplast, Tandem mass spectrometry}, doi = {10.1002/elps.201900452}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1002/elps.201900452}, author = {Cheng, Jianhui and Morin, Gregg B. and Chen, David D. Y.} } @article {MA201829, title = {Characterization of plant polysaccharides from Dendrobium officinale by multiple chromatographic and mass spectrometric techniques}, journal = {Journal of Chromatography A}, volume = {1547}, year = {2018}, pages = {29 - 36}, keywords = {Liquid chromatography, POLYSACCHARIDES, Tandem mass spectrometry}, issn = {0021-9673}, doi = {https://doi.org/10.1016/j.chroma.2018.03.006}, url = {http://www.sciencedirect.com/science/article/pii/S0021967318302784}, author = {Huiying Ma and Keke Zhang and Qing Jiang and Diya Dai and Hongli Li and Wentao Bi and David Da Yong Chen} } @article {2663, title = {A Carboxy-Terminal Affinity Tag for the Purification and Mass Spectrometric Characterization of Integral Membrane Proteins}, journal = {Journal of Proteome Research}, volume = {8}, number = {5}, year = {2009}, note = {ISI Document Delivery No.: 441BWTimes Cited: 3Cited Reference Count: 22Wong, Julie P. Reboul, Emmanuelle Molday, Robert S. Kast, Juergen}, month = {May}, pages = {2388-2396}, type = {Article}, abstract = {G-protein-coupled receptors (GPCRs) and other structurally and functionally related membrane proteins represent particularly attractive targets for drug discovery. Integral membrane proteins are often difficult to purify from native contexts, and lack of sufficient quantities hampers subsequent structural and functional proteomic studies. We describe here an optimized enrichment strategy involving a membrane protein-compatible 1D4 affinity tag that is derived from the carboxy-terminal nine amino residues of bovine rhodopsin, and its corresponding tag-specific, high-affinity monoclonal antibody. When two GPCRs as well as two related ATP binding cassette (ABC) transporters are expressed in their functional forms in human cell lines, we have shown that a single detergent and wash condition can be employed for the purification of all said membrane proteins. Subsequent in-gel digestion with trypsin and mass spectrometric peptide analysis resulted in high sequence coverage for the ABC transporters ABCA1-1D4 and ABCA4-1D4. In contrast, digestion by various enzymatic combinations was necessary to obtain the best sequence coverage for affinity-enriched GPCRs CXCR4-1D4 and CCR5-1D4 as compared against other entries in an annotated spectrum library. Furthermore, specific enzyme combinations were necessary to produce suitable peptides for deducing N-glycosylation sites on CXCR4. Our results demonstrate that the 1D4-tag enrichment strategy is a versatile tool for the characterization of integral membrane proteins that can be employed for functional proteomic studies.}, keywords = {1D4 affinity tag, 2-DIMENSIONAL, annotated spectrum libraries, ATP binding cassette transporters, BINDING, ELECTROPHORESIS, EXPRESSION, G-protein-coupled, membrane protein enrichment, N-LINKED GLYCOSYLATION, PROTEOMICS, proteotypic peptides, RECEPTOR, RECEPTORS, RHODOPSIN, ROD OUTER SEGMENTS, SITES, SYNTHETIC PEPTIDES, Tandem mass spectrometry}, isbn = {1535-3893}, url = {