Glycosylinositol Phospholipid Anchors of the Scrapie and Cellular Prion Proteins Contain Sialic Acid

Stahl, Neil; Baldwin, Michael A.; Hecker, Rolf; Pan, Keh-Ming; Burlingame, Alma L.; Prusiner, Stanley B.

doi:10.1021/bi00136a600

Cited by 274 publications

(228 citation statements)

References 79 publications

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“…We would predict Man 4 -GPIs to be most prevalent in brain and colorectal tissues where hSMP3 expression is greatest, and this notion is supported by structural data indicating that Man 4 -GPIs are present on various GPI-anchored brain proteins (6,26). Additionally, hSMP3 expression is barely detectable in the spleen and peripheral blood leukocytes, suggesting that Man 3 -GPIs may predominate in these tissues.…”

Section: Discussionmentioning

confidence: 66%

“…In support of this notion, a fourth mannose is present on 7 of 10 characterized mammalian protein-bound GPIs (6, 7, 26 -31, 41, 42). Six of the 7 Man 4 -GPIs were isolated from proteins purified directly from primary sources like homogenized organ tissue (6,26,28,29,31) or urine (27). Only 1 was from a protein purified from a cultured mouse cell line (30).…”

Section: Discussionmentioning

confidence: 99%

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Human Smp3p Adds a Fourth Mannose to Yeast and Human Glycosylphosphatidylinositol Precursors in Vivo

Taron

Colussi

Wiedman³

et al. 2004

Journal of Biological Chemistry

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Yeast and human glycosylphosphatidylinositol (GPI) precursors differ in the extent to which a fourth mannose is present as a side branch of the third core mannose. A fourth mannose addition to GPIs has scarcely been detected in studies of mammalian GPI synthesis but is an essential step in the Saccharomyces cerevisiae pathway. We report that human SMP3 encodes a functional homolog of the yeast Smp3 GPI fourth mannosyltransferase. Expression of hSMP3 in yeast complements growth and biochemical defects of smp3 mutants and permits in vivo mannosylation of trimannosyl (Man 3 )-GPIs. Immunolocalization shows that hSmp3p resides in the endoplasmic reticulum in human cells. Northern analysis of mRNA from human tissues and cell lines indicates that hSMP3 is expressed in most tissues, with the highest levels in brain and colon, but its mRNA is nearly absent from cultured human cell lines. Correspondingly, increasing expression of hSMP3 in cultured HeLa cells causes abundant formation of three putative tetramannosyl (Man 4 )-GPIs. Our data indicate that hSmp3p functions as a mannosyltransferase that adds a fourth mannose to certain Man 3 -GPIs during biosynthesis of the human GPI precursor, and suggest it may do so in a tissue-specific manner.Glycosylphosphatidylinositols (GPIs) 1 are essential glycolipids synthesized by all eukaryotes. Many GPIs become covalently attached to the carboxyl termini of various secretory proteins and serve to anchor them to the exterior face of the plasma membrane (1, 2). Others remain protein-free and are distributed in the membranes of major cellular organelles and the plasma membrane (3, 4). GPIs are synthesized in the endoplasmic reticulum (ER) by stepwise addition of components to phosphatidylinositol. The end product of GPI synthesis is a "complete precursor" (5) that is substrate for the GPI transamidase complex that attaches it to proteins. Many of the steps and enzymes of GPI precursor assembly are conserved between humans and Saccharomyces cerevisiae and produce precursors with a common core structure of EthN-PO 4 -6Man␣1,2Man␣1,6Man␣1,4GlcN␣1,6Ins-PO 4 -lipid. In both pathways, the glycan portion of the GPI may be modified further with side branching ethanolamine phosphate (EthN-P) moieties on the first and second mannoses (6 -19).A notable difference in GPI structure between yeast and mammals is the extent to which a fourth mannose (Man-4) is present as a ␣1,2-linked side branch of the third mannose (Man-3). In S. cerevisiae, late stage intermediates in GPI precursor synthesis (17, 19 -21), the presumed GPI transamidase substrates (5), and protein-bound GPIs (22) all contain four mannoses. Addition of Man-4 to trimannosyl-GPIs (Man 3 -GPIs) by the essential Smp3 mannosyltransferase is a mandatory step in yeast GPI precursor assembly which precedes the addition of the terminal EthN-P to Man-3 through which the GPI is ultimately attached to protein (23). Thus, it is probable that all yeast GPI transamidase substrates bear four mannoses. Conversely, studies of the synthesis of mammal...

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Section: Discussionmentioning

confidence: 66%

Section: Discussionmentioning

confidence: 99%

Human Smp3p Adds a Fourth Mannose to Yeast and Human Glycosylphosphatidylinositol Precursors in Vivo

Taron

Colussi

Wiedman³

et al. 2004

Journal of Biological Chemistry

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“…Accumulation of PrP Sc in the CNS leads to prion diseases, a family of transmissible neurodegenerative maladies that are 100% lethal (11). PrP C is posttranslationally modified with two N-linked glycans and a glycosylphosphatidylinositol (GPI) anchor that is attached to the C-terminal residue Ser-230 (the residue number is provided for mouse PrP C ) (12)(13)(14)(15)(16). Upon conversion of PrP C into PrP Sc , the N-linked glycans and GPI are carried over, giving rise to glycosylated and GPI-anchored PrP Sc (17,18).…”

Section: Prions or Prpmentioning

confidence: 99%

Sialylation of Glycosylphosphatidylinositol (GPI) Anchors of Mammalian Prions Is Regulated in a Host-, Tissue-, and Cell-specific Manner

Katorcha

Srivastava

Klimova

et al. 2016

Journal of Biological Chemistry

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Prions or PrPSc are proteinaceous infectious agents that consist of misfolded, self-replicating states of the prion protein or PrP C . PrP C is posttranslationally modified with N-linked glycans and a sialylated glycosylphosphatidylinositol (GPI) anchor. Sc . Remarkably, the proportion of sialo-versus asialo-GPIs was found to be controlled by host, tissue, and cell type but not prion strain. In summary, this study found no strain-specific preferences for selecting PrP C with sialo-versus asialo-GPIs. Instead, this work suggests that the sialylation status of GPIs within PrP Sc is regulated in a cell-, tissue-, or host-specific manner and is likely to be determined by the specifics of GPI biosynthesis.

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“…Post translational modifications include two Asn-linked glycosylation sites [14,15], a GPI anchor [21], a single disulphide bond [22], and partial proline hydroxylation near the N-terminus [23]. The glycosylation of the prion protein has been suggested to play a fundamental role in the transmissible spongiform encephalopathies (TSEs) [24 -26].…”

Section: Prion Protein (Prp Sc )mentioning

confidence: 99%

Precursor ion scanning for detection and structural characterization of heterogeneous glycopeptide mixtures

Ritchie

Gill

Deery

et al. 2002

J. Am. Soc. Mass Spectrom.

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The structure of N-linked glycans is determined by a complex, anabolic, intracellular pathway but the exact role of individual glycans is not always clear. Characterization of carbohydrates attached to glycoproteins is essential to aid understanding of this complex area of biology. Specific mass spectral detection of glycopeptides from protein digests may be achieved by on-line HPLC-MS, with selected ion monitoring (SIM) for diagnostic product ions generated by cone voltage fragmentation, or by precursor ion scanning for terminal saccharide product ions, which can yield the same information more rapidly. When glycosylation is heterogeneous, however, these approaches can result in spectra that are complex and poorly resolved. We have developed methodology, based around precursor ion scanning for ions of high m/z, that allows site specific detection and structural characterization of glycans at high sensitivity and resolution. These methods have been developed using the standard glycoprotein, fetuin, and subsequently applied to the analysis of the N-linked glycans attached to the scrapieassociated prion protein, PrP Sc . These glycans are highly heterogeneous and over 30 structures have been identified and characterized site specifically. Product ion spectra have been obtained on many glycopeptides confirming structure assignments. The glycans are highly fucosylated and carry Lewis X or sialyl Lewis X epitopes and the structures are in-line with previous results. [Abbreviations: Hex-Hexose, C 6 H 12 O 6 carbohydrates, including mannnose and galactose; HexNAc-N-acetylhexosamine, C 8 H 15 NO 6 carbohydrates, including N-acetylglucosamine and N-acetylgalactosamine; GlcNAc-N-acetylglucosamine; GalNAc-N-acetylgalactosamine; Fuc-Fucose; NeuAC-N-acetylneuraminic acid or sialic acid; TSE-Transmissible Spongiform Encephalopathy.] (J Am Soc Mass Spectrom 2002Spectrom , 13, 1065Spectrom -1077

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Glycosylinositol Phospholipid Anchors of the Scrapie and Cellular Prion Proteins Contain Sialic Acid

Cited by 274 publications

References 79 publications

Human Smp3p Adds a Fourth Mannose to Yeast and Human Glycosylphosphatidylinositol Precursors in Vivo

Human Smp3p Adds a Fourth Mannose to Yeast and Human Glycosylphosphatidylinositol Precursors in Vivo

Sialylation of Glycosylphosphatidylinositol (GPI) Anchors of Mammalian Prions Is Regulated in a Host-, Tissue-, and Cell-specific Manner

Precursor ion scanning for detection and structural characterization of heterogeneous glycopeptide mixtures

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