glycolipids and gpi anchors glycolipids and gpi anchors glycolipids and gpi anchors glycolipids and...

Glycolipids and GPI anchorsGlycolipids and GPI anchorsGlycolipids and GPI anchorsGlycolipids and GPI anchors

• Learn the structures, and some functions of the most Learn the structures, and some functions of the most abundant vertebrate glycolipidsabundant vertebrate glycolipids

• Glucosylceramide and the skin barrier• Galactosylceramide & sulfatide in myelin• Glycolipids in cell-cell interactions and membrane receptor

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• Learn the structures, distribution and biosynthesis of Learn the structures, distribution and biosynthesis of GPI anchorsGPI anchors

• GPI-anchored proteins and their anchor structures in people and parasites

• GPI anchor biosynthesis and en bloc transfer

• Glycolipids and GPI anchors in diseaseGlycolipids and GPI anchors in disease• Glycolipids and lysosomal storage diseases• GPI anchors in anemia



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ObjectivesObjectivesObjectivesObjectives

http://www.youtube.com/watch?v=v9j-g0qspdk


Major membrane lipid classes

PLC

GLYCEROLIPID




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Most vertebrate glycolipids are Most vertebrate glycolipids are glycosphingolipidsglycosphingolipids

Most vertebrate glycolipids are Most vertebrate glycolipids are glycosphingolipidsglycosphingolipids


Glucosylceramide is a key intermediate in skin Glucosylceramide is a key intermediate in skin barrier formationbarrier formation


Skin water barrier. Formation and secretion of lamellar bodies (LB) at stratum granulosum – stratum corneum Skin water barrier. Formation and secretion of lamellar bodies (LB) at stratum granulosum – stratum corneum interface. Precursor lipids, including glucosylceramide, sphingomyelin, glycerophospholipids, and cholesterol interface. Precursor lipids, including glucosylceramide, sphingomyelin, glycerophospholipids, and cholesterol sulfate, are packaged into LB in the upper epidermis. Fusion of the LB with the apical plasma membrane in the sulfate, are packaged into LB in the upper epidermis. Fusion of the LB with the apical plasma membrane in the uppermost layer of the epidermis (stratum granulosum), allows extrusion of lipid precursors into the extracellular uppermost layer of the epidermis (stratum granulosum), allows extrusion of lipid precursors into the extracellular domain. Enzymatic processing of precursor lipids generates the major lipid classes required for epidermal barrier domain. Enzymatic processing of precursor lipids generates the major lipid classes required for epidermal barrier function.function.

Holleran et al (2006) Febs Letters 23:5456


Normal mouse skinNormal mouse skin

““Gaucher” Gaucher” mouse skin mouse skin

(glucocerebrosidase -/-)(glucocerebrosidase -/-)

Holleran et al (1994) J Clin Invest 93:1756




Normal human skinNormal human skin

Human type 2Human type 2Gaucher disease skin Gaucher disease skin (glucocerebrosidase- (glucocerebrosidase-

deficient)deficient)

Holleran et al (1994) J Clin Invest 93:1756







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galactosylceramide and it’s 3-sulfated from, galactosylceramide and it’s 3-sulfated from, sulfatide, are major lipids in the brainsulfatide, are major lipids in the brain

galactosylceramide and it’s 3-sulfated from, galactosylceramide and it’s 3-sulfated from, sulfatide, are major lipids in the brainsulfatide, are major lipids in the brain

Together, GalCer and sulfatide comprise up to 1.9% of brain fresh weight!


galactosylceramide & sulfatide are the major galactosylceramide & sulfatide are the major glycans in the brain, and major lipids in myelinglycans in the brain, and major lipids in myelingalactosylceramide & sulfatide are the major galactosylceramide & sulfatide are the major

glycans in the brain, and major lipids in myelinglycans in the brain, and major lipids in myelin

Myelin total lipids - mole percent

Mo

le r

atio

0

10

20

30

40Phospho-

lipids

Choles-terol

Galacto-lipids

Glycan ratios in the brain


Delcomyn, F. (1998) Foundations of Neurobiology.

W.H. Freeman, New York.

Mutant mice lacking GalCer synthetase (Mutant mice lacking GalCer synthetase (Ugt8aUgt8a) ) oror GalCer 3-sulfotransferase ( GalCer 3-sulfotransferase (Gal3st1) Gal3st1) display myelin display myelin

defects and associated behavioral deficitsdefects and associated behavioral deficits

Mutant mice lacking GalCer synthetase (Mutant mice lacking GalCer synthetase (Ugt8aUgt8a) ) oror GalCer 3-sulfotransferase ( GalCer 3-sulfotransferase (Gal3st1) Gal3st1) display myelin display myelin

defects and associated behavioral deficitsdefects and associated behavioral deficits

Honke et al (2002) PNAS 99:4227

Honke et al (2002) PNAS 99:4227

Popko (2000) Glia 29:149


Glycoglycerolipids are not widely distributed in vertebrates, but Glycoglycerolipids are not widely distributed in vertebrates, but 3-O-sulfated galactosyldiacylglycerol (seminolipid) makes up 3-O-sulfated galactosyldiacylglycerol (seminolipid) makes up 90% of the glycolipid in mammalian testes. 90% of the glycolipid in mammalian testes. Gal3st1Gal3st1-null males -null males are infertile due to an early block in spermatogenesis.are infertile due to an early block in spermatogenesis.

Glycoglycerolipids are not widely distributed in vertebrates, but Glycoglycerolipids are not widely distributed in vertebrates, but 3-O-sulfated galactosyldiacylglycerol (seminolipid) makes up 3-O-sulfated galactosyldiacylglycerol (seminolipid) makes up 90% of the glycolipid in mammalian testes. 90% of the glycolipid in mammalian testes. Gal3st1Gal3st1-null males -null males are infertile due to an early block in spermatogenesis.are infertile due to an early block in spermatogenesis.





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GlcCer based glycolipid families:GlcCer based glycolipid families:-All share a lactosylceramide (LacCer) coreAll share a lactosylceramide (LacCer) core-Nomenclature is based on the next sugars in the neutral tetrasaccharide Nomenclature is based on the next sugars in the neutral tetrasaccharide corecore-Classified as “neutral glycosphingolipids”, “sulfated glycosphingolipids” or Classified as “neutral glycosphingolipids”, “sulfated glycosphingolipids” or “gangliosides” (sialylated glycosphingolipids)“gangliosides” (sialylated glycosphingolipids)-Nomenclature describes the length of the core and any substituents, Nomenclature describes the length of the core and any substituents, number the sugars from the reducing (Cer) end out by roman numeralsnumber the sugars from the reducing (Cer) end out by roman numerals-Biosynthesized stepwise by individual glycosphingolipids, some of which Biosynthesized stepwise by individual glycosphingolipids, some of which are glycolipid-specific, others of which also make glycoprotein glycansare glycolipid-specific, others of which also make glycoprotein glycans


Subfamily series Structure Abbreviation

Lacto Galβ1-3GlcNAcβ1-3Galβ1-4GlcβCer Lc4Cer

Neolacto Galβ1-4GlcNAcβ1-3Galβ1-4GlcβCer nLc4Cer

Ganglio Galβ1-3GalNAcβ1-4Galβ1-4GlcβCer Gg4Cer

Globo GalNAcβ1-3Galα1-4Galβ1-4GlcβCer Gb4Cer

Lactosylceramide (LacCer)


http://www.functionalglycomics.org/fg/

http://www.genome.jp/kegg/pathway.html

NeuAc 3 Gal 3 GalNAc 4 (NeuAc 8 NeuAc 3) Gal 4 Glc Cer

Ganglioside GT1bGanglioside GT1b

Gal GalNAc

Gal GlcNeuAc

NeuAc

NeuAc

Gangliosides are embedded in the extracellular leaflet of the Gangliosides are embedded in the extracellular leaflet of the plasma membrane with their glycans extending outwardplasma membrane with their glycans extending outward

DeMarco & Woods (2009) Glycobiology 19, 344DeMarco & Woods (2009) Glycobiology 19, 344

Rat cerebellar granule cell axons in culture immunostained for ganglioside GD1aRat cerebellar granule cell axons in culture immunostained for ganglioside GD1a

HIC (phase) image overlay imageanti-GD1a mAb

Gangliosides are on ALL vertebrate cells, but are the Gangliosides are on ALL vertebrate cells, but are the dominant glycan structures on nerve cellsdominant glycan structures on nerve cells

* non-neuronal cells

The same four structures comprise the great majority The same four structures comprise the great majority of gangliosides in the brains of all mammalsof gangliosides in the brains of all mammals

GM1Cer

GD1bCer

GT1bCer

GD1aCer

(28%)

(26%)

(25%)

(18%)

Glc Gal GalNAc NeuAc

Values are for human; Gangliosides shown constitute 97% of total brain gangliosidesTettamanti et al. (1973) Biochim Biophys Acta 296, 160

Brain ganglioside biosynthesisBrain ganglioside biosynthesis

ST3GAL5ST3GAL5:: “Infantile-onset symptomatic epilepsy syndrome caused by a homozygous “Infantile-onset symptomatic epilepsy syndrome caused by a homozygous loss-of-loss-of-

functionfunction mutation of GM3 synthase” mutation of GM3 synthase” Simpson MA et al. Nature Genet 36, 1225 (2004)Simpson MA et al. Nature Genet 36, 1225 (2004)

B4GALNT1B4GALNT1:: “Alteration of ganglioside biosynthesis responsible for complex hereditary spastic “Alteration of ganglioside biosynthesis responsible for complex hereditary spastic paraplegia” paraplegia” Boukhris et al. Am J Human Genet 93, 118 (2013)Boukhris et al. Am J Human Genet 93, 118 (2013)

Concepts of glycolipid function: Concepts of glycolipid function: ciscis and and transtransConcepts of glycolipid function: Concepts of glycolipid function: ciscis and and transtrans

ciscis: regulation by : regulation by lateral associationlateral associationciscis: regulation by : regulation by lateral associationlateral association

transtrans: regulation by : regulation by cell-cell recognition cell-cell recognition transtrans: regulation by : regulation by cell-cell recognition cell-cell recognition

Note: Because of their long unsaturated lipid chains, glycolipids cluster together, Note: Because of their long unsaturated lipid chains, glycolipids cluster together, along with cholesterol, selected other lipids, and selected proteins, in “lipid rafts”along with cholesterol, selected other lipids, and selected proteins, in “lipid rafts”Note: Because of their long unsaturated lipid chains, glycolipids cluster together, Note: Because of their long unsaturated lipid chains, glycolipids cluster together, along with cholesterol, selected other lipids, and selected proteins, in “lipid rafts”along with cholesterol, selected other lipids, and selected proteins, in “lipid rafts”

Regina Todeschini & Hakomori (2008) Biochim Biophys Acta 1780:421





Glycolipid purification and analysis: Glycolipid purification and analysis: Solvent extract, partition/chromatography, TLCSolvent extract, partition/chromatography, TLCGlycolipid purification and analysis: Glycolipid purification and analysis: Solvent extract, partition/chromatography, TLCSolvent extract, partition/chromatography, TLC

• Homogenize cells or Homogenize cells or tissuestissues

• Add methanol and Add methanol and chloroformchloroform

• Remove precipitated Remove precipitated protein and DNA by protein and DNA by centrifugationcentrifugation

• Isolate glycolipids by Isolate glycolipids by partition and/or partition and/or adsorptive adsorptive chromatographychromatography

• Analyze by TLC Analyze by TLC

Mouse brain gangliosides – silica TLC – resorcinol sialic acid stain

Mouse sialyltransferase mutants

origin -




Subfamily series Structure Abbreviation

Lacto Galβ1-3GlcNAcβ1-3Galβ1-4GlcβCer Lc4Cer

Neolacto Galβ1-4GlcNAcβ1-3Galβ1-4GlcβCer nLc4Cer

Ganglio Galβ1-3GalNAcβ1-4Galβ1-4GlcβCer Gg4Cer

Globo GalNAcβ1-3Galα1-4Galβ1-4GlcβCer Gb4Cer

Lactosylceramide (LacCer)


http://www.functionalglycomics.org/fg/

http://www.genome.jp/kegg/pathway.html

Example: Total neolactoseries monsialylated glycolipids from Example: Total neolactoseries monsialylated glycolipids from human neutrophils – diversity within a structural themehuman neutrophils – diversity within a structural themeExample: Total neolactoseries monsialylated glycolipids from Example: Total neolactoseries monsialylated glycolipids from human neutrophils – diversity within a structural themehuman neutrophils – diversity within a structural theme

Nimrichter et al (2008) Blood, 112:3744


Neolactoseries monsialylated glycolipids on human neutrophilsNeolactoseries monsialylated glycolipids on human neutrophilsNeolactoseries monsialylated glycolipids on human neutrophilsNeolactoseries monsialylated glycolipids on human neutrophils

Nimrichter et al (2008) Blood,

112:3744


Separation of human neutrophil gangliosides by HPLCSeparation of human neutrophil gangliosides by HPLCSeparation of human neutrophil gangliosides by HPLCSeparation of human neutrophil gangliosides by HPLC

Nimrichter et al (2008) Blood,

112:3744


Concepts of glycolipid function: Concepts of glycolipid function: transtrans recognition in health and disease recognition in health and diseaseConcepts of glycolipid function: Concepts of glycolipid function:

transtrans recognition in health and disease recognition in health and disease

• Bacterial toxins bind to glycolipidsBacterial toxins bind to glycolipids– Shigatoxin (verotoxin) binds to the neutral glycosphingolipid Shigatoxin (verotoxin) binds to the neutral glycosphingolipid

GbGb33 (Gal (Galα1-α1-4Gal4Galβ1-4β1-4GlcGlcβ1-β1-Cer)Cer)

– Cholera and E. Coli enterotoxins bind to the ganglioside Cholera and E. Coli enterotoxins bind to the ganglioside GM1 (GalGM1 (Galβ1-3β1-3GalNAcGalNAcβ1-4(β1-4(NeuAcNeuAcα2-3)α2-3)GalGalβ1-4β1-4GlcGlcβ1-β1-Cer)Cer)

– Tetanus and botulinum neurotoxins toxins bind to nervous Tetanus and botulinum neurotoxins toxins bind to nervous system gangliosides with different specificitiessystem gangliosides with different specificities

• A brain lectin, myelin-associated glycoprotein (MAG), A brain lectin, myelin-associated glycoprotein (MAG), binds to gangliosides GD1a and GT1b to meidate binds to gangliosides GD1a and GT1b to meidate axon-myelin interactions*axon-myelin interactions*

• An endothelial cell lectin, E-selectin, binds to An endothelial cell lectin, E-selectin, binds to sialylated fucosylated glycolipids on human sialylated fucosylated glycolipids on human neutrophils to initiate inflammation*neutrophils to initiate inflammation*

* These lectins will be discussed in a later lecture* These lectins will be discussed in a later lecture


Research correlation: Multivalent toxin inhibitor Research correlation: Multivalent toxin inhibitor mimics glycolipid in the cell membrane mimics glycolipid in the cell membrane


LEFT: ABLEFT: AB55 bacterial toxins (cholera toxin shown) have bacterial toxins (cholera toxin shown) have

5 glycolipid cell surface binding subunits surrounding a 5 glycolipid cell surface binding subunits surrounding a “payload” toxic subunit (not shown) that is inserted into “payload” toxic subunit (not shown) that is inserted into the cell only after binding.the cell only after binding.

LEFT: ABLEFT: AB55 bacterial toxins (cholera toxin shown) have bacterial toxins (cholera toxin shown) have

5 glycolipid cell surface binding subunits surrounding a 5 glycolipid cell surface binding subunits surrounding a “payload” toxic subunit (not shown) that is inserted into “payload” toxic subunit (not shown) that is inserted into the cell only after binding.the cell only after binding.

Merritt et al. (1994) Protein Sci 3:166

RIGHT: Taking RIGHT: Taking advantage of the ABadvantage of the AB55

structure of structure of vertotoxin, and its vertotoxin, and its GbGb33 specific binding, specific binding,

Kitov et al Kitov et al synthesized a synthesized a “starfish” shaped “starfish” shaped inhibitor with five inhibitor with five arms, each carrying arms, each carrying two copies of Gbtwo copies of Gb33. .

RIGHT: Taking RIGHT: Taking advantage of the ABadvantage of the AB55

structure of structure of vertotoxin, and its vertotoxin, and its GbGb33 specific binding, specific binding,

Kitov et al Kitov et al synthesized a synthesized a “starfish” shaped “starfish” shaped inhibitor with five inhibitor with five arms, each carrying arms, each carrying two copies of Gbtwo copies of Gb33. .

Kitov et al. (2000) Nature 403:669






ABOVE: “Starfish”GbABOVE: “Starfish”Gb33 blocks verotoxin binding at sub-nM blocks verotoxin binding at sub-nM

concentrations – a million times more potent than Gbconcentrations – a million times more potent than Gb33

ABOVE: “Starfish”GbABOVE: “Starfish”Gb33 blocks verotoxin binding at sub-nM blocks verotoxin binding at sub-nM

concentrations – a million times more potent than Gbconcentrations – a million times more potent than Gb33



Right: “Starfish”GbRight: “Starfish”Gb33 crystallized with verotoxin – crystallized with verotoxin –

a perfect pentomeric sandwicha perfect pentomeric sandwichRight: “Starfish”GbRight: “Starfish”Gb33 crystallized with verotoxin – crystallized with verotoxin –

a perfect pentomeric sandwicha perfect pentomeric sandwich







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A small but diverse group of proteins are attached to A small but diverse group of proteins are attached to the cell surface by a “glycosylphosphatidylinositol” the cell surface by a “glycosylphosphatidylinositol” (GPI) anchor(GPI) anchor

A small but diverse group of proteins are attached to A small but diverse group of proteins are attached to the cell surface by a “glycosylphosphatidylinositol” the cell surface by a “glycosylphosphatidylinositol” (GPI) anchor(GPI) anchor

GPI anchors have a minimal (core) structure (black font) with variable additional glycan, fatty acid ester, and phosphoethanolamine groups (blue font)

Taylor and Drikamer (2011) Introduction to Glycobiology


Essentials of Glycobiology Second Edition

GPI anchors are GPI anchors are complex & diversecomplex & diverseGPI anchors are GPI anchors are complex & diversecomplex & diverse

• Glucosamine with Glucosamine with free amine – rarefree amine – rare

• Phosphodiesters at Phosphodiesters at each endeach end

• Diverse Diverse modifications along modifications along the corethe core

R3


A group of select proteins are GPI anchored*A group of select proteins are GPI anchored*A group of select proteins are GPI anchored*A group of select proteins are GPI anchored*MammalsMammals

Erythrocyte CD59 and decay Erythrocyte CD59 and decay acceleration factor (DAF)acceleration factor (DAF) complement regulationcomplement regulation

Alkaline phosphataseAlkaline phosphatase cell-surface hydrolasecell-surface hydrolase

5′-Nucleotidase5′-Nucleotidase cell-surface hydrolasecell-surface hydrolase

Renal dipeptidaseRenal dipeptidase cell-surface hydrolasecell-surface hydrolase

TrehalaseTrehalase cell-surface hydrolasecell-surface hydrolase

Neural cell adhesion Neural cell adhesion molecule 120 (NCAM-120)molecule 120 (NCAM-120) adhesion moleculeadhesion molecule

Neural cell adhesion Neural cell adhesion molecule TAG-1molecule TAG-1 adhesion moleculeadhesion molecule

CD58CD58 adhesion moleculeadhesion molecule

FcFcγγIII receptorIII receptor Fc receptorFc receptor

Ciliary neurotrophic factor Ciliary neurotrophic factor receptor (CNTFR) α subunitreceptor (CNTFR) α subunit neural receptorneural receptor

Glial-cell-derived Glial-cell-derived neurotrophic factor receptor neurotrophic factor receptor (GDNFR) α subunit(GDNFR) α subunit

neural receptorneural receptor

CD14CD14 LPS receptorLPS receptor

Prion protein (PrP)Prion protein (PrP) unknownunknown

Glypican family of GPI-Glypican family of GPI-anchored proteoglycansanchored proteoglycans

extracellular matrix extracellular matrix componentcomponent

Trypanosoma bruceTrypanosoma brucei i variant surface variant surface glycoprotein (VSG)glycoprotein (VSG)

protective coatprotective coat

Leishmania majorLeishmania major promastigote surface promastigote surface protease (PSP)protease (PSP)

bound complement bound complement degradationdegradation

Trypanosoma cruziTrypanosoma cruzi GPI- GPI-anchored mucinsanchored mucins host cell invasionhost cell invasion

Plasmodium falciparumPlasmodium falciparum merozoite surface protein 1 merozoite surface protein 1 (MSP-1)(MSP-1)

erythrocyte invasionerythrocyte invasion

Toxoplasma gondiiToxoplasma gondii surface surface antigen 1 (SAG-1)antigen 1 (SAG-1) host cell invasionhost cell invasion

Entamoeba histolyticaEntamoeba histolytica GPI GPI proteophosphoglycansproteophosphoglycans virulence factorvirulence factor

ParasitesParasites

*plus some in yeast, plants, some other species*plus some in yeast, plants, some other species


Variety is the spice of… GPI anchorsVariety is the spice of… GPI anchorsVariety is the spice of… GPI anchorsVariety is the spice of… GPI anchors






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Preassembled GPI Preassembled GPI anchors are covalently anchors are covalently transferred transferred en blocen bloc via via amide linkage to an amide linkage to an amino acid near the amino acid near the carboxy terminus of a carboxy terminus of a nacient protein, at the nacient protein, at the same time releasing same time releasing the C-terminal protein the C-terminal protein fragment. This occurs fragment. This occurs in the ER.in the ER.

Although there is no Although there is no “consensus sequence,” “consensus sequence,” per se, for GPI transfer, per se, for GPI transfer, likely sites are identified likely sites are identified by their structural by their structural features.features.

Preassembled GPI Preassembled GPI anchors are covalently anchors are covalently transferred transferred en blocen bloc via via amide linkage to an amide linkage to an amino acid near the amino acid near the carboxy terminus of a carboxy terminus of a nacient protein, at the nacient protein, at the same time releasing same time releasing the C-terminal protein the C-terminal protein fragment. This occurs fragment. This occurs in the ER.in the ER.

Although there is no Although there is no “consensus sequence,” “consensus sequence,” per se, for GPI transfer, per se, for GPI transfer, likely sites are identified likely sites are identified by their structural by their structural features.features.


Like N-linked glycan biosynthesis, the precursor starts assembly on the cytoplasmic Like N-linked glycan biosynthesis, the precursor starts assembly on the cytoplasmic face of the ER, then is flipped inside for further processing. GlcNAc transfer and face of the ER, then is flipped inside for further processing. GlcNAc transfer and deacetylation, inositol fatty acid esterification, and multiple ethanolamine phosphate deacetylation, inositol fatty acid esterification, and multiple ethanolamine phosphate group transfers by separate enzymes are noteworthy.group transfers by separate enzymes are noteworthy.

Like N-linked glycan biosynthesis, the precursor starts assembly on the cytoplasmic Like N-linked glycan biosynthesis, the precursor starts assembly on the cytoplasmic face of the ER, then is flipped inside for further processing. GlcNAc transfer and face of the ER, then is flipped inside for further processing. GlcNAc transfer and deacetylation, inositol fatty acid esterification, and multiple ethanolamine phosphate deacetylation, inositol fatty acid esterification, and multiple ethanolamine phosphate group transfers by separate enzymes are noteworthy.group transfers by separate enzymes are noteworthy.


GPI anchor synthesis has been studied most extensively in parasites, which express GPI anchor synthesis has been studied most extensively in parasites, which express millions of GPI-anchored proteins and glycans on the surface of each cell. millions of GPI-anchored proteins and glycans on the surface of each cell. GPI anchor synthesis has been studied most extensively in parasites, which express GPI anchor synthesis has been studied most extensively in parasites, which express millions of GPI-anchored proteins and glycans on the surface of each cell. millions of GPI-anchored proteins and glycans on the surface of each cell.

http://www.lifesci.dundee.ac.uk/people/mike-ferguson

African sleeping sickness Chagas diseaseleishmaniasis

GPI anchor synthesis in parasites is complex with some interesting twists, such as GPI anchor synthesis in parasites is complex with some interesting twists, such as fatty acid remodeling. Since GPI anchored coat proteins are virulence factors, they fatty acid remodeling. Since GPI anchored coat proteins are virulence factors, they are being exploited as targets for therapy.are being exploited as targets for therapy.

GPI anchor synthesis in parasites is complex with some interesting twists, such as GPI anchor synthesis in parasites is complex with some interesting twists, such as fatty acid remodeling. Since GPI anchored coat proteins are virulence factors, they fatty acid remodeling. Since GPI anchored coat proteins are virulence factors, they are being exploited as targets for therapy.are being exploited as targets for therapy.

Essentials of Glycobiology

Second Edition

To study the structures and functions of GPI anchored proteins, they can be To study the structures and functions of GPI anchored proteins, they can be released under biologically compatible conditions using bacterial released under biologically compatible conditions using bacterial phosphatidylinositol-specific phospholipase C (PIPLC), or decomposed specifically phosphatidylinositol-specific phospholipase C (PIPLC), or decomposed specifically by nitrous acid, which cleaves at the glucosamine residue forming a 2,5-by nitrous acid, which cleaves at the glucosamine residue forming a 2,5-anhydromannose residue.anhydromannose residue.

To study the structures and functions of GPI anchored proteins, they can be To study the structures and functions of GPI anchored proteins, they can be released under biologically compatible conditions using bacterial released under biologically compatible conditions using bacterial phosphatidylinositol-specific phospholipase C (PIPLC), or decomposed specifically phosphatidylinositol-specific phospholipase C (PIPLC), or decomposed specifically by nitrous acid, which cleaves at the glucosamine residue forming a 2,5-by nitrous acid, which cleaves at the glucosamine residue forming a 2,5-anhydromannose residue.anhydromannose residue.





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Clinical correlation: defects in glycolipid degradationClinical correlation: defects in glycolipid degradationClinical correlation: defects in glycolipid degradationClinical correlation: defects in glycolipid degradation

Disease name Disease name (glycolipid)(glycolipid) Enzyme or protein deficiencyEnzyme or protein deficiency Clinical symptomsClinical symptoms

Tay–Sachs (GM2)Tay–Sachs (GM2) β-β-hexosaminidase Ahexosaminidase A neurodegeneration, death by 4 yearsneurodegeneration, death by 4 years

Sandhoff (GM2)Sandhoff (GM2) β-β-hexosaminidase A and Bhexosaminidase A and B neurodegeneration, death by 4 yearsneurodegeneration, death by 4 years

GM1 gangliosidosis GM1 gangliosidosis (GM1)(GM1) β-β-galactosidasegalactosidase progressive neurological disease and skeletal progressive neurological disease and skeletal

dysplasia in severe infantile formdysplasia in severe infantile form

Fabry*Fabry*(Gb3)(Gb3) α-α-galactosidasegalactosidase

severe pain, angiokeratoma, corneal severe pain, angiokeratoma, corneal opacities, death from renal or cerebrovascular opacities, death from renal or cerebrovascular diseasedisease

Gaucher’s*Gaucher’s*(GlcCer)(GlcCer) β-β-glucoceramidaseglucoceramidase hepatosplenomegaly hepatosplenomegaly

neurodegeneration (severe form only)neurodegeneration (severe form only)

KrabbeKrabbe(GalCer)(GalCer) β-β-galactoceramidasegalactoceramidase early onset with progression to severe mental early onset with progression to severe mental

and motor deteriorationand motor deterioration

Metachromatic Metachromatic leukodystrophy leukodystrophy

(sulfatide)(sulfatide)

arylsulfatase A arylsulfatase A (cerebroside sulfatase)(cerebroside sulfatase)

mental regression, peripheral neuropathy, mental regression, peripheral neuropathy, seizures, dementiaseizures, dementia

Saposin deficiencySaposin deficiency(many)(many) saposin precursorsaposin precursor neurodegeneration, death by 4 yearsneurodegeneration, death by 4 years

*enzyme replacement therapy available*enzyme replacement therapy availableEssentials of Glycobiology

Second Edition

Gaucher disease – cause, symptoms and treatmentGaucher disease – cause, symptoms and treatmentGaucher disease – cause, symptoms and treatmentGaucher disease – cause, symptoms and treatment

Hepatosplenomegaly: Grossly enlarged liver and spleen

http://www.gauchercare.com/

The cure: enzyme replacement therapy

Andersson et al.. (2008) Pediatrics 122:1182

A new direction – substrate reduction therapyA new direction – substrate reduction therapyA new direction – substrate reduction therapyA new direction – substrate reduction therapy

n-butyl-deoxynojirimycin = Miglustat = Zavesca®




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Clinical correlation: Paroxysmal nocturnal hemoglobinuriaClinical correlation: Paroxysmal nocturnal hemoglobinuriaClinical correlation: Paroxysmal nocturnal hemoglobinuriaClinical correlation: Paroxysmal nocturnal hemoglobinuria

http://www.bioch.ox.ac.uk/glycob/archive/

CD59, a GPI-anchored “complement defense protein” terminal complement complex

http://trialx.com/

The cause, somatic mutation of the PIG-A gene resulting in failure to transfer GlcNAc to phosphatidylinositol, and loss of multiple GPI-anchored complement control proteins.

Essentials of Glycobiology

Clinical correlation: Paroxysmal nocturnal hemoglobinuriaClinical correlation: Paroxysmal nocturnal hemoglobinuriaClinical correlation: Paroxysmal nocturnal hemoglobinuriaClinical correlation: Paroxysmal nocturnal hemoglobinuria

Kelly R J et al. Blood (2011)117:6786

Anti-complement protein 5 treatment improves outcomesAnti-complement protein 5 treatment improves outcomesAnti-complement protein 5 treatment improves outcomesAnti-complement protein 5 treatment improves outcomes

per year per year




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Documents

anchor structures

gpi anchorslearn

cellcell interactions

en bloc transferglycolipids

apical plasma membrane

epidermal barrier function

epidermis stratum granulosum

j clin