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http://purl.uniprot.org/citations/36213313http://www.w3.org/1999/02/22-rdf-syntax-ns#typehttp://purl.uniprot.org/core/Journal_Citation
http://purl.uniprot.org/citations/36213313http://www.w3.org/1999/02/22-rdf-syntax-ns#typehttp://purl.uniprot.org/core/Journal_Citation
http://purl.uniprot.org/citations/36213313http://www.w3.org/2000/01/rdf-schema#comment"Chondroitin sulfate proteoglycans (CSPGs) are extracellular matrix components composed of linear glycosaminoglycan (GAG) side chains attached to a core protein. CSPGs play a vital role in neurodevelopment, signal transduction, cellular proliferation and differentiation and tumor metastasis through interaction with growth factors and signaling proteins. These pleiotropic functions of proteoglycans are regulated spatiotemporally by the GAG chains attached to the core protein. There are over 70 chondroitin sulfate-linked proteoglycans reported in cells, cerebrospinal fluid and urine. A core glycan linker of 3-6 monosaccharides attached to specific serine residues can be extended by 20-200 disaccharide repeating units making intact CSPGs very large and impractical to analyze. The current paradigm of CSPG analysis involves digesting the GAG chains by chondroitinase enzymes and analyzing either the protein part, the disaccharide repeats, or both by mass spectrometry. This method, however, provides no information about the site of attachment or the composition of linker oligosaccharides and the degree of sulfation and/or phosphorylation. Further, the analysis by mass spectrometry and subsequent identification of novel CSPGs is hampered by technical challenges in their isolation, less optimal ionization and data analysis. Unknown identity of the linker oligosaccharide also makes it more difficult to identify the glycan composition using database searching approaches. Following chondroitinase digestion of long GAG chains linked to tryptic peptides, we identified intact GAG-linked peptides in clinically relevant samples including plasma, urine and dermal fibroblasts. These intact glycopeptides including their core linker glycans were identified by mass spectrometry using optimized stepped higher energy collision dissociation and electron-transfer/higher energy collision dissociation combined with hybrid database search/de novo glycan composition search. We identified 25 CSPGs including three novel CSPGs that have not been described earlier. Our findings demonstrate the utility of combining enrichment strategies and optimized high-resolution mass spectrometry analysis including alternative fragmentation methods for the characterization of CSPGs.

Supplementary information

The online version contains supplementary material available at 10.1007/s42485-022-00092-3."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.org/dc/terms/identifier"doi:10.1007/s42485-022-00092-3"xsd:string
http://purl.uniprot.org/citations/36213313http://purl.org/dc/terms/identifier"doi:10.1007/s42485-022-00092-3"xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/author"Pandey A."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/author"Pandey A."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/author"Raymond K."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/author"Raymond K."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/author"Budhraja R."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/author"Budhraja R."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/author"Garapati K."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/author"Garapati K."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/author"Ramarajan M.G."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/author"Ramarajan M.G."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/author"Saraswat M."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/author"Saraswat M."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/date"2022"xsd:gYear
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/date"2022"xsd:gYear
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/name"J. Proteins Proteom."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/name"J. Proteins Proteom."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/pages"187-203"xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/pages"187-203"xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/title"Mass spectrometric analysis of chondroitin sulfate-linked peptides."xsd:string
http://purl.uniprot.org/citations/36213313http://purl.uniprot.org/core/title"Mass spectrometric analysis of chondroitin sulfate-linked peptides."xsd:string
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