Analysis of congenital disorder of glycosylation-Id in a yeast model system shows diverse site-specific under-glycosylation of glycoproteins

Bailey, Ulla-Maja, Jamaluddin, Muhammad Fairuz and Schulz, Benjamin L. (2012) Analysis of congenital disorder of glycosylation-Id in a yeast model system shows diverse site-specific under-glycosylation of glycoproteins. Journal of Proteome Research, 11 11: 5376-5383. doi:10.1021/pr300599f

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Author Bailey, Ulla-Maja
Jamaluddin, Muhammad Fairuz
Schulz, Benjamin L.
Title Analysis of congenital disorder of glycosylation-Id in a yeast model system shows diverse site-specific under-glycosylation of glycoproteins
Journal name Journal of Proteome Research   Check publisher's open access policy
ISSN 1535-3893
1535-3907
Publication date 2012-10-16
Sub-type Article (original research)
DOI 10.1021/pr300599f
Volume 11
Issue 11
Start page 5376
End page 5383
Total pages 18
Place of publication Washington, DC, United States
Publisher American Chemical Society
Collection year 2013
Language eng
Formatted abstract Asparagine-linked glycosylation is a common post-translational modification of proteins in eukaryotes.  Mutations in the human ALG3 gene cause changed levels and altered glycan structures on mature glycoproteins and are the cause of a severe congenital disorder of glycosylation (CDG-Id). Diverse glycoproteins are also under-glycosylated in Saccharomyces cerevisae alg3 mutants. Here we analyzed sitespecific glycosylation occupancy in this yeast model system using peptide-N-glycosidase F to label glycosylation sites with an asparagine-aspartate conversion that creates a new endoproteinase AspN cleavage site, followed by proteolytic digestion, and detection of peptides and glycopeptides by LC−ESI−MS/MS. We used this analytical method to identify and measure site-specific glycosylation occupancy in alg3 mutant and wild type yeast strains. We found decreased site-specific N-glycosylation occupancy in the alg3 knockout strain preferentially at Asn-Xaa-Ser sequences located in secondary structural elements, features previously associated with poor glycosylation efficiency.  Furthermore, we identified 26 previously experimentally unverified glycosylation sites. Our results provide insights into the underlying mechanisms of disease in CDG-Id, and our methodology will be useful in site-specific glycosylation analysis in many model systems and clinical applications.
Keyword Oligosaccharyltransferase
Mass spectrometry
Glycosylation occupancy
ALG
alg3
CDG
N-glycosylation
PNGase F
Open Access Mandate Compliance Yes - Open Access (Author post-print in repository)
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ
Additional Notes epub ahead of print

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2013 Collection
School of Chemistry and Molecular Biosciences
 
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Citation counts: TR Web of Science Citation Count  Cited 10 times in Thomson Reuters Web of Science Article | Citations
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Created: Fri, 19 Oct 2012, 10:22:17 EST by Mrs Louise Nimwegen on behalf of School of Chemistry & Molecular Biosciences