Dataset Information

Extensive hypoglycosylation of serum N-glycoproteins in SRD5A3 deficiency

ABSTRACT: Polyprenol reductase is an enzyme encoded by the SRD5A3 gene, which catalyzes the synthesis of dolichol from polyprenols. Dolichol serves as a carrier for glycan precursors in N-linked glycosylation or monosaccharides in O-glycosylation, C-mannosylation, and GPI anchor biosynthesis. Pathogenic variants in SRD5A3 can result in a congenital disorder of glycosylation (CDG), SRD5A3-CDG, which is inherited in an autosomal recessive manner. Most serum proteins undergo glycosylation and changes in the glycosylation levels of numerous serum glycoproteins have been associated with pathological consequences. Despite the critical role of SRD5A3 in glycosylation, the impact of its deficiency on the glycosylation of serum proteins remains largely unexplored. In this study, we used tandem mass tags (TMT)-based multiplexed quantitation approach to analyze serum N-glycoproteins and proteins in SRD5A3-CDG patients and controls. We quantified 2,200 serum N-glycopeptides from 359 N-glycosites from 204 serum proteins. Extensive hypoglycosylation of serum proteins was observed in patients, with 245 of 291 dysregulated glycopeptides showing hypoglycosylation. These significantly changing glycopeptides belonged to several known abundant serum glycoproteins including haptoglobin, plasma serine protease inhibitor, alpha-1-B glycoprotein, alpha-2-macroglobulin and ceruloplasmin. We also detected changes in glycopeptides from albumin which has recently been shown to be glycosylated. Overall, our study provides novel insights into alterations in the glycosylation status of serum proteins in SRD5A3-CDG that could facilitate further investigations into diagnostics and therapeutics aspects of this CDG.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Blood Serum

SUBMITTER: Akhilesh Pandey

LAB HEAD: Akhilesh Pandey

PROVIDER: PXD053534 | Pride | 2026-06-08

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	AJ_SRD5A3-CDG_Serum_TMT_NGP_B10_B11_F5.raw	Raw
	AJ_SRD5A3-CDG_Serum_TMT_NGP_B12_C1_F6.raw	Raw
	AJ_SRD5A3-CDG_Serum_TMT_NGP_B2_B3_F1.raw	Raw
	AJ_SRD5A3-CDG_Serum_TMT_NGP_B4_B5_F2.raw	Raw
	AJ_SRD5A3-CDG_Serum_TMT_NGP_B6_B7_F3.raw	Raw

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Publications

Albumin as a glycoprotein biomarker in congenital disorders of glycosylation.

Garapati Kishore K Jain Anu A Joshi Neha N Sachdeva Gunveen S GS Nam Dowoon D Saraswat Mayank M Pasupuleti Raghavendra Rao RR Schultz Matthew J MJ Kozicz Tamas T Morava Eva E Pandey Akhilesh A

Molecular genetics and metabolism 20260210 4

Congenital disorders of glycosylation (CDG) are rare inherited disorders resulting from defects in cellular glycosylation machinery. Albumin has recently been shown to be N-glycosylated at two non-canonical glycosylation sites. We applied multiplexed mass spectrometry-based glycoproteomics to identify site-specific N-glycosylation alterations in albumin from patients with PMM2-CDG, MPI-CDG, SRD5A3-CDG, MAN1B1-CDG and PGM1-CDG. Our findings demonstrate that the glycosylation of albumin is indeed ...[more]

PMID: 41713138

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Project description:Phosphomannomutase 2 (PMM2) converts mannose-6-phospahate to mannose-1-phosphate; the substrate for GDP-mannose, a building block of the glycosylation biosynthetic pathway. Pathogenic variants in the PMM2 gene have been shown to be associated with protein hypoglycosylation causing PMM2-congenital disorder of glycosylation (PMM2-CDG). While mannose supplementation improves glycosylation in vitro, but not in vivo, we hypothesized that liposomal delivery of mannose-1-phosphate could increase the stability and delivery of the activated sugar to enter the targeted compartments of cells. Thus, we studied the effect of liposome-encapsulated mannose-1-P (GLM101) on global protein glycosylation and on the cellular proteome in skin fibroblasts from individuals with PMM2-CDG, as well as in individuals with two N-glycosylation defects early in the pathway, namely ALG2-CDG and ALG11-CDG. We leveraged multiplexed proteomics and N-glycoproteomics in fibroblasts derived from different individuals with various pathogenic variants in PMM2, ALG2 and ALG11 genes. Proteomics data revealed a moderate but significant change in the abundance of some of the proteins in all CDG fibroblasts upon GLM101 treatment. On the other hand, N-glycoproteomics revealed the GLM101 treatment enhanced the expression levels of several high-mannose and complex/hybrid glycopeptides from numerous cellular proteins in individuals with defects in PMM2 and ALG2 genes. Both PMM2-CDG and ALG2-CDG exhibited several-fold increase in glycopeptides bearing Man6 and higher glycans and a decrease in Man5 and smaller glycan moieties, suggesting that GLM101 helps in the formation of mature glycoforms. These changes in protein glycosylation were observed in all individuals irrespective of their genetic variants. ALG11-CDG fibroblasts also showed increase in high mannose glycopeptides upon treatment; however, the improvement was not as dramatic as the other two CDG. Overall, our findings suggest that treatment with GLM101 overcomes the genetic block in the glycosylation pathway and can be used as a potential therapy for CDG with enzymatic defects in early steps in protein N-glycosylation.

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.

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