Dataset Information

Serial In-solution digestion protocol for mass spectrometry-based glycomics and proteomics analysis

ABSTRACT: Advancement in mass spectrometry has revolutionised the field of proteomics. However, there remains a gap in the analysis of protein post-translational modifications (PTMs), particularly for glycosylation. Glycosylation, the most common form of PTM, is involved in most biological processes; thus, analysis of glycans along with proteins is crucial to answering important biologically relevant questions. Of particular interest is the brain extracellular matrix (ECM), which has been called the “final frontier” in neuroscience that consists of highly glycosylated proteins. Among these, proteoglycans (PGs) contain large glycan structures called glycosaminoglycans (GAGs) and form crucial ECM components, including perineuronal nets (PNNs), shown to be altered in neuropsychiatric diseases. Thus, there is a growing need for high-throughput methods that combine GAG (glycomics) and PGs (proteomics) analysis to unravel the complete biological picture. The protocol presented here integrates glycomics and proteomics to analyze multiple classes of biomolecules.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Bos Taurus (bovine) Homo Sapiens (human) Mus Musculus (mouse)

TISSUE(S): Brain, Blood Plasma, Articular Cartilage

SUBMITTER: Manveen Sethi

LAB HEAD: Prof. Joseph Zaia

PROVIDER: PXD017513 | Pride | 2020-04-30

REPOSITORIES: pride

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Serial in-solution digestion protocol for mass spectrometry-based glycomics and proteomics analysis.

Sethi Manveen K MK Downs Margaret M Zaia Joseph J

Molecular omics 20200420 4

Advancement in mass spectrometry has revolutionized the field of proteomics. However, there remains a gap in the analysis of protein post-translational modifications (PTMs), particularly for glycosylation. Glycosylation, the most common form of PTM, is involved in most biological processes; thus, analysis of glycans along with proteins is crucial to answering important biologically relevant questions. Of particular interest is the brain extracellular matrix (ECM), which has been called the "fina ...[more]

PMID: 32309832

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Project description:Glioblastoma (GBM) is the most common and malignant primary brain tumor. The extracellular matrix (ECM), also known as the matrisome, helps determine glioma invasion, adhesion, and growth. Little attention, however, has been paid to glycosylation of the ECM components that constitute the majority of glycosylated protein mass and presumed biological properties. To acquire a comprehensive understanding of the biological functions of the matrisome and its components, including proteoglycans and glycosaminoglycans (GAGs), in GBM tumorigenesis, and to identify potential biomarker candidates, we studied the alterations of GAGs, including heparan sulfate (HS) and chondroitin sulfate (CS), the core proteins of proteoglycans, and other glycosylated matrisomal proteins in GBM subtypes vs. control human brain tissue samples. We scrutinized the proteomics data to acquire in-depth site-specific glycoproteomic profiles of the GBM subtypes that will assist in identifying specific glycosylation changes in GBM. We observed an increase in CS 6-O sulfation and a decrease in HS 6-O sulfation, accompanied by an increase in unsulfated CS and HS disaccharides in GBM vs. control samples. Several core matrisome proteins, including proteoglycans (decorin, biglycan, agrin, prolargin, glypican-1, CSPG4), tenascin, fibronectin, hyaluronan link protein 1 and 2, laminins, and collagens, were differentially regulated in GBM vs. controls. Interestingly, a higher degree of collagen hydroxyprolination was also observed for GBM vs. controls. Further, two proteoglycans, CSPG4 and agrin, were significantly lower, about 6–fold for IDH-mutant compared to the WT GBM samples. Differential regulation of O-glycopeptides for proteoglycans, including brevican, neurocan, and versican, was observed for GBM subtypes vs. controls. Moreover, an increase in levels of glycosyltransferase and glycosidase enzymes was observed for GBM when compared to control samples. We also report distinct protein, peptide, and glycopeptide features for GBM subtypes comparisons. Taken together, our study informs understanding of the alterations to key matrisomal molecules that occur during GBM development.

			Action	DRS
	AGP_batch01_01.mzML	Mzml
	AGP_batch01_01.mzid.gz	Mzid
	AGP_batch01_01.raw	Raw
	AGP_batch01_02.mzML	Mzml
	AGP_batch01_02.mzid.gz	Mzid