Dataset Information

Sialic Acid Linkage-Specific Alkylamidation-Based Method for Comprehensive N- and O-Glycoproteomic Profiling of Human Plasma

ABSTRACT: Sialic acids are attached to glycoproteins via alpha2,3- or alpha2,6-linkages and play important roles in disease pathogenesis. Sialyl linkage-specific derivatization of intact N- and/or O-glycopeptides remains challenging because of the diverse reactive functional groups within peptides. In this report, we present a comprehensive workflow for identifying and quantifying sialylated N- and O-glycopeptides that integrates dimethyl labeling for amino-group blocking, sialic acid linkage-specific alkylamidation, and label-free quantification.

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Yuri Miura

PROVIDER: PXD080569 | JPOST Repository | Wed Jun 10 00:00:00 BST 2026

REPOSITORIES: jPOST

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Dataset's files

Source:

			Action	DRS
	Data%20list_SALSA-Peptide.xlsx	Xlsx
	PD2.4-Byonic3.10%20(hPlasma_N-Glycan).zip	Other
	PD2.4-Byonic3.10%20(hPlasma_O-Glycan).zip	Other
	PD2.4-Byonic3.8%20(hAGP_N-Glycan).zip	Other
	PD2.4-Byonic3.8%20(hFET_N+O-Glycan).zip	Other

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Publications

Sialic Acid Linkage-Specific Alkylamidation-Based Method for N- and O-Glycoproteomic Profiling of Human Plasma.

Tsumoto Hiroki H Nishikaze Takashi T Sekiya Sadanori S Umezawa Keitaro K Kawakami Kyojiro K Masui Yukie Y Ikebe Kazunori K Kamide Kei K Hirata Takumi T Gondo Yasuyuki Y Iwamoto Shinichi S Tanaka Koichi K Miura Yuri Y

Analytical chemistry 20260708 28

Sialic acids are attached to glycoproteins via α2,3- or α2,6-linkages and play important roles in disease pathogenesis. Therefore, accurate discrimination and quantification of these linkage isomers are essential. Although several chemical derivatization strategies to discriminate the isomers by mass differences have been developed for the analysis of released N-glycans, sialyl linkage-specific derivatization of intact N- and/or O-glycopeptides remains challenging because of ...[more]

PMID: 42421262

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Project description:Human noroviruses, globally the main cause of viral gastroenteritis, show strain specific affinity for histo-blood group antigens (HBGA) and can successfully be propagated ex vivo in human intestinal enteroids (HIEs). HIEs established from jejunal stem cells of individuals with different ABO, Lewis and secretor geno- and phenotypes, show varying susceptibility to such infections. Using bottom-up glycoproteomic approaches we have defined and compared the N-linked glycans of glycoproteins of seven jejunal HIEs. Membrane proteins were extracted, trypsin digested, and glycopeptides enriched by hydrophilic interaction liquid chromatography and analyzed by nanoLC-MS/MS. The Byonic software was used for glycopeptide identification followed by hands-on verifications and interpretations. Glycan structures and attachment sites were identified from MS 2 spectra obtained by higher-energy collision dissociation through analysis of diagnostic saccharide oxonium ions (B-ions), stepwise glycosidic fragmentation of the glycans (Y-ions), and peptide sequence ions (b- and y-ions). Altogether 694 unique glycopeptides from 93 glycoproteins were identified. The N-glycans encompassed pauci- and oligomannose, hybrid- and complex-type structures. Notably, polyfucosylated HBGA-containing glycopeptides of the four glycoproteins tetraspanin-8, carcinoembryonic antigen-related cell adhesion molecule 5, sucrose-isomaltase and aminopeptidase N were especially prominent and were characterized in detail and related to donor ABO, Lewis and secretor types of each HIE. Virtually no sialylated N-glycans were identified for these glycoproteins suggesting that terminal sialylation was infrequent compared to fucosylation and HBGA biosynthesis. This approach gives unique site-specific information on the structural complexity of N-linked glycans of glycoproteins of human HIEs and provides a platform for future studies on the role of host glycoproteins in gastrointestinal infectious diseases.

Project description:SRD5A3-CDG is a congenital disorder of glycosylation (CDG) resulting from pathogenic variants in SRD5A3 and follows an autosomal recessive inheritance pattern. The enzyme encoded by SRD5A3, polyprenal reductase, plays a crucial role in synthesizing lipid precursors essential for N-linked glycosylation. Despite insights from functional studies into its enzymatic function, there remains a gap in understanding global changes in patient cells. We sought to identify glycoproteomic and proteomic signatures specific to SRD5A3-CDG, potentially aiding in biomarker discovery and advancing our understanding of disease mechanisms. Using tandem mass tag (TMT)-based relative quantitation, we analyzed fibroblasts derived from five patients along with control fibroblasts. Glycoproteomics analysis by liquid chromatographyâ€“tandem mass spectrometry (LC-MS/MS) identified 3,047 glycopeptides with 544 unique glycosylation sites from 276 glycoproteins. Of these, 418 glycopeptides showed statistically significant changes with 379 glycopeptides decreased (p<0.05) in SRD5A3-CDG patient-derived samples. These included high mannose, complex and hybrid glycan-bearing glycopeptides. High mannose glycopeptides from protocadherin Fat 4 and integrin alpha-11 and complex glycopeptides from CD55 were among the most significantly decreased glycopeptides. Proteomics analysis led to the identification of 5,933 proteins, of which 873 proteins showed statistically significant changes. Decreased proteins included cell surface glycoproteins, various mitochondrial protein populations and proteins involved in the N-glycosylation pathway. Lysosomal proteins such as N-acetylglucosamine-6-sulfatase and procathepsin-L also showed reduced levels of phosphorylated mannose-containing glycopeptides. Our findings point to disruptions in glycosylation pathways as well as energy metabolism and lysosomal functions in SRD5A3-CDG, providing clues to improved understanding and management of patients with this disorder.

Dataset Information

Sialic Acid Linkage-Specific Alkylamidation-Based Method for Comprehensive N- and O-Glycoproteomic Profiling of Human Plasma

Dataset's files

Publications

Sialic Acid Linkage-Specific Alkylamidation-Based Method for <i>N</i>- and <i>O</i>-Glycoproteomic Profiling of Human Plasma.

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Dataset Information

Sialic Acid Linkage-Specific Alkylamidation-Based Method for Comprehensive N- and O-Glycoproteomic Profiling of Human Plasma

Dataset's files

Publications

Sialic Acid Linkage-Specific Alkylamidation-Based Method for &lt;i&gt;N&lt;/i&gt;- and &lt;i&gt;O&lt;/i&gt;-Glycoproteomic Profiling of Human Plasma.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets

Sialic Acid Linkage-Specific Alkylamidation-Based Method for <i>N</i>- and <i>O</i>-Glycoproteomic Profiling of Human Plasma.