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• W2111667172 abstract "Yeast glycan biosynthetic pathways are commonly studied through metabolic incorporation of an exogenous radiolabeled compound into a target glycan. In Saccharomyces cerevisiae glycosylphosphatidylinositol (GPI) biosynthesis, [(3) H]inositol has been widely used to identify intermediates that accumulate in conditional GPI synthesis mutants. However, this approach also labels non-GPI lipid species that overwhelm detection of early GPI intermediates during chromatography. In this study, we show that despite lacking the ability to metabolize N-acetylglucosamine (GlcNAc), S. cerevisiae is capable of importing low levels of extracellular GlcNAc via almost all members of the hexose transporter family. Furthermore, expression of a heterologous GlcNAc kinase gene permits efficient incorporation of exogenous [(14) C]GlcNAc into nascent GPI structures in vivo, dramatically lowering the background signal from non-GPI lipids. Utilizing this new method with several conditional GPI biosynthesis mutants, we observed and characterized novel accumulating lipids that were not previously visible using [(3) H]inositol labeling. Chemical and enzymatic treatments of these lipids indicated that each is a GPI intermediate likely having one to three mannoses and lacking ethanolamine phosphate (Etn-P) side-branches. Our data support a model of yeast GPI synthesis that bifurcates after the addition of the first mannose and that includes a novel branch that produces GPI species lacking Etn-P side-branches." @default.
• W2111667172 created "2016-06-24" @default.
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• W2111667172 date "2012-01-18" @default.
• W2111667172 modified "2023-09-24" @default.
• W2111667172 title "Uptake of radiolabeled GlcNAc into Saccharomyces cerevisiae via native hexose transporters and its in vivo incorporation into GPI precursors in cells expressing heterologous GlcNAc kinase" @default.
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• W2111667172 doi "https://doi.org/10.1111/j.1567-1364.2011.00778.x" @default.
• W2111667172 hasPubMedCentralId "https://www.ncbi.nlm.nih.gov/pmc/articles/3498731" @default.
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