Protein glycosylation is a post-translational modification (PTM) responsible for many aspects of proteomic diversity and biological regulation. Assignment of intact glycan structures to specific protein attachment sites is a critical step towards elucidating the function encoded in the glycome. Previously, we developed isotope-targeted glycoproteomics (IsoTaG) as a mass-independent mass spectrometry method to characterize azide-labeled intact glycopeptides from complex proteomes. Here, we extend the IsoTaG approach with the use of alkynyl sugars as metabolic labels and employ new probes in analysis of the sialylated glycoproteome from PC-3 cells. Using an Orbitrap Fusion Tribrid mass spectrometer, we identified 699 intact glycopeptides from 192 glycoproteins. These intact glycopeptides represent a total of eight sialylated glycan structures across 126 N- and 576 O-glycopeptides. IsoTaG is therefore an effective platform for identification of intact glycopeptides labeled by alkynyl or azido sugars and will facilitate further studies of the glycoproteome.
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http://dx.doi.org/10.1007/s00216-016-9934-9 | DOI Listing |
Anal Chem
January 2025
College of Chemistry, Jilin University, Changchun 130012, China.
Smart-responsive materials have attracted much attention in the enrichment of post-translational modifications of proteins. In this work, for the first time, we developed a smart enrichment strategy (MNPs-l-DOPA/PEI-SP) based on the change in hydrophilic properties of spiropyran under the regulation of light and pH to realize the controllable enrichment and release of intact glycopeptides. The enrichment mechanism and possible binding mechanism were verified by theoretical calculations.
View Article and Find Full Text PDFInt J Biol Macromol
December 2024
Institute of Mass Spectrometry, Zhejiang Engineering Research Center of Advanced Mass Spectrometry and Clinical Application, School of Material Science and Chemical Engineering, Ningbo University, Ningbo 315211, China; Zhenhai Institute of Mass Spectrometry, Ningbo 315211, China. Electronic address:
N-glycosylation is crucial in the process of wheat yellow mosaic virus (WYMV) infection, but changes in site-specific N-glycosylation of proteins during WYMV infection have not been well studied. In this study, we employed an intact glycopeptide approach to analyze mock- and WYMV-infected wheat plants. We found that most glycoproteins have N-glycans containing paucimannose or complex/hybrid chains.
View Article and Find Full Text PDFFood Chem
December 2024
Chinese Academy of Inspection and Quarantine, Beijing 100176, People's Republic of China. Electronic address:
Glycoproteins, which are involved in numerous biological functions, are among the most critical functional ingredients in an edible bird's nest (EBN). To gain a comprehensive understanding of the glycoprotein species within EBN, a label-free, site-specific glycoproteomic approach was used to analyze their N-glycoproteins, N-glycopeptides, and N-glycans systematically. A total of 127 N-glycoproteins were identified in EBN, of which 72 were found in house-EBN and 63 in cave-EBN, yielding 4195 and 5649 glycopeptides, respectively.
View Article and Find Full Text PDFMethods Mol Biol
December 2024
Department of Biochemistry, Center for Biomedical Mass Spectrometry, Boston University School of Medicine, Boston, MA, USA.
Mass spectrometry-based investigation of the heterogeneous glycoproteome from complex biological specimens is a robust approach to mapping the structure, function, and dynamics of the glycome and proteome. Sampling whole wet tissues often provides a large amount of starting material; however, there is a reasonable variability in tissue handling prior to downstream processing steps, and it is difficult to capture all the different biomolecules from a specific region. The on-slide tissue digestion approach, outlined in this protocol chapter, is a simple and cost-effective method that allows comprehensive mapping of the glycoproteome from a single spot of tissue of 1 mm or greater diameter.
View Article and Find Full Text PDFAnal Bioanal Chem
December 2024
XJTLU Wisdom Lake Academy of Pharmacy-BEAVER Biomedical Joint Laboratory, Suzhou, 215123, China.
Protein glycosylation is one of the most important post-translational modifications, implicated in the development of various diseases, including neurodegenerative diseases, diabetes, and cancers. However, the low content of glycoproteins in biological samples, the diversity and heterogeneity of glycan structures, and insensitive detection methods make glycosylation analysis challenging. As a result, efficient enrichment of glycopeptides from complex samples is a critical step.
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