essentials of glycobiology march 30th, 2004 ajit varki
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Essentials of Glycobiology March 30th, 2004 Ajit Varki. Lecture 1 Course Overview & General Introduction. Online at NCBI http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=Books. Lecture 1 - Key Points. What is Glycobiology? Central Dogma Basic definitions - PowerPoint PPT PresentationTRANSCRIPT
Essentials of Glycobiology
March 30th, 2004
Ajit Varki
Lecture 1 Course Overview & General Introduction
Online at NCBI http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=Books
Lecture 1 - Key Points
• What is Glycobiology?
• Central Dogma
• Basic definitions
• Monosaccharides - the structural units of glycans
• Oligoaccharides consist of monosaccharides linked together
• Nomenclature and symbolic representations
• Major classes of glycoconjugates and oligosaccharides
• Clustered oligosaccharides
• Proteins can bind at either the ends of glycans or within the chains
DNA RNA PROTEIN CELL ORGANISM
DNA
ORGANISM
? ?
All Cells Are Coated with “Glycans”
Electron micrograph of a human lymphocyte (Ruthenium Red staining)
LIPIDS
DNA RNA PROTEINS
MATRIXCELL
GLYCOPROTEINSPROTEOGLYCANSGLYCOLIPIDS
SIGNALLINGMOLECULES
TISSUES & ORGANS
TRANSCRIPTION FACTORS
PHYSICAL ENVIRONMENT
MICROBESPARASITES
ENZYMES
DIET
DNA
SUGARS
ORGANISM
DNA
Macromolecules
Macromolecule Building Block AproximateMass
PossibleVariations in a
TrimerProtein Amino acids 125 ’ 104-105 6
Nucleic Acid Nucleotides 330 ’ 103-109 6
Lipid Fatty acids 250 ’ 103 NA
Carbohydrate Monosaccharides 200 ’ 102-106 1,056 to 27,648!
What is Glycobiology?
¶ Study of the structure, biosynthesis and biology of saccharides (sugar chains or “glycans”) in nature.
¶ Glycobiology is an integrative science, crossing all subfields of chemistry, biology and medicine.
¶ Chemical analysis and biochemistry dominated the field in the early part of the century.
¶ In the 1960’s, glycans in the extracellular matrix, on the cell surface, and in the interior of cells were found to have biological properties independent of the underlying protein or lipid.
¶ The term “Glycobiology” was coined in 1988 by Rademacher, Parekh, and Dwek: Annu Rev Biochem. 57:785-838.
“Central Dogma”
Glycans occur in patterns that are characteristic of the cell-type, tissue and organism
Glycan patterning is not template driven
Instead, the glycan composition and pattern depends on expression of biosynthetic enzyme (transferases), substrate specificity, and the availability of precursors (i.e., more like “assembly-line driven)
Lecture 1 - Key Points
• What is Glycobiology?
• Central Dogma
• Basic definitions
• Monosaccharides - the structural units of glycans
• Oligoaccharides consist of monosaccharides linked together
• Nomenclature and symbolic representations
• Major classes of glycoconjugates and oligosaccharides
• Clustered oligosaccharides
• Proteins can bind at either the ends of glycans or within the chains
Basic Definitions
• Monosaccharide: A carbohydrate that cannot be hydrolyzed into a simpler carbohydrate. The building block of oligosaccharides and polysaccharides.
• Oligosaccharide: Linear or branched chain of monosaccharides attached to one another via glycosidic linkages. The number of monosaccharide units can vary.
• Polysaccharide: Glycan composed of repeating monosaccharides, generally greater than ten monosaccharide units in length.
• Carbohydrate, glycan, saccharide, sugar: Generic terms used interchangeably. Includes monosaccharides, oligosaccharides, polysaccharides, and derivatives of these compounds. Carbohydrates consist of “hydrated carbon”, [CH2O]n
• Preferred generic term is “Glycan”
Monosaccharides - the basic structural unit
• Carbonyl group at the end of the carbon chain (aldoses) or at an inner carbon (ketoses) has potential reducing power. This end is called the reducing terminus, or reducing end
• The ring form of a monosaccharide generates a chiral (anomeric) center (at C-1 for aldo sugars or at C-2 for keto sugars). Notice that other positions are chiral, which therefore imparts stereochemical information
HO
CHO
H
HHO
OHH
OHH
CH2OH
HOO
H
H
HO
OH
OHHH
H
OH
H O
OH
H
OH
OH
HH
OH
CH2OH
H
1
2
3
4
5
6
1
23
4
5
6
123
4
5
6
Fischer Hayworth Stereochemical
Oligosaccharides and Polysaccharides
-linkage -linkage
OH O
H
H
H
OHH
OH
CH2OH
H1
23
4
5
6
H O
H
H
OHH
OH
CH2OH
H1
23
4
5
6
O
H O H
OH
OH
HH
OH
CH2OH
H1
23
4
5
6
O
Glycosidic linkage can be or The convention is to draw the reducing end to the right and the non-reducing end to the left. Once in glycosidic linkage, ring can no longer open Reducing end is still called the same even when it is covelantly bound to another macromolecule (tying up its reducing power)
Common Monosaccharides
NeutralSugars
AminoSugars
AcidicSugars
OOH
OH
OH
HO
CH2OH
Galactose(Gal)
OOH
OHOHHO
CH2OH
Mannose(Man)
OOH
OH
OHHO
Xylose(Xyl)
OOH
OH
OH
HO
CH3
Fucose(Fuc)
OOH
OH
OHHO
CH2OH
Glucose(Glc)
CHOHCH2OH
CHOHO
COOH
OH
AcN
OH
Sialic acids(Sia)
OOH
OH
OHHO
COOH
Glucuronic acid(GlcA)
OOH
OH
OHHO
COOH
Iduronic acid(IdoA)
OOH
NAC
OH
HOCH2OH
N-acetyl Galactosamine(GalNAc)
OOH
NAC
OHHO
CH2OH
N-acetyl Glucosamine(GlcNAc)
Lecture 1 - Key Points
• What is Glycobiology?
• Central Dogma
• Basic definitions
• Monosaccharides - the structural units of glycans
• Oligoaccharides consist of monosaccharides linked together
• Nomenclature and symbolic representations
• Major classes of glycoconjugates and oligosaccharides
• Clustered oligosaccharides
• Proteins can bind at either the ends of glycans or within the chains
Symbolic Representation of Common Monosaccharides
To simplify the structural complexity of glycans, we will generally use a standard set of symbols to represent sugars.
= Glucose (Glc)= Mannose (Man)= Galactose (Gal)= N-acetylglucosamine (GlcNAc)= N-acetylgalactosamine (GalNAc)= N-acetylhexosamine, unspecified (HexNAc)
= Fucose (Fuc)= Xylose (Xyl)= Sialic acid, unspecified (Sia)= Glucuronic acid (GlcA)= Iduronic acid (IdoA)= Uronic acid, unspecified (HexA)
Hexose,unspecified (Hex)
Symbol set is being modified for second edition of “Essentials”, in a consensus agreement with NCBI, Consortium for Functional Glycomics, KEGG and other national and international bodies
Saccharide Modifications increase the diversity and functionality of glycans
= GlcNAc= GlcA= Gal= Xyl = IdoA
NS NSNS NSNS NSNS2S 2S2S 2S6S 6S 6S 6S 6S6S
3S
Ac = O-acetyl P = Phosphate S = O-Sulfate NS = N-Sulfate NH2 = free amino group
Lecture 1 - Key Points
• What is Glycobiology?
• Central Dogma
• Basic definitions
• Monosaccharides - the structural units of glycans
• Oligoaccharides consist of monosaccharides linked together
• Nomenclature and symbolic representations
• Major classes of glycoconjugates and oligosaccharides
• Clustered oligosaccharides
• Proteins can bind at either the ends of glycans or within the chains
Glycoconjugates
Glycoconjugate: A compound in which one or more glycans (the glycone) are covalently linked to a non-carbohydrate moiety (the aglycone).
Glycoproteins: A protein with one or more covalently bound glycans.
Glycolipids: A molecule containing a saccharide linked to a lipid.
Proteoglycans: Any glycoprotein with one or more covalently attached glycosaminoglycan chains.
N-glycan N-glycanGPI-glycan
GPI-acyl chains
Polypeptide
Schematic representation of the Thy-1 glycoprotein
Major Classes of Animal Glyconconjugates
N-GlcNAc linkedoligosaccharideN-linked chain
N-glycan
O-GalNAc linkedoligosaccharideO-linked chain
O-glycan
O-Xyl linkedoligosaccharide
GlycosaminoglycanGAG chain
Ceramide-linkedoligosaccharides
GlycosphingolipidsGlycolipids
Glycophospholipidanchor
GPI anchorPI-glycan
O--linked GlcNAcO-GlcNAc
OSer/Thr
4
NAsn
4
63
OSer
4
3
3
4
NH2
6
4
2
OSer/Thr
OUTSIDE
INSIDE
*
*Several additional kinds of O-glycans have emerged recently
Major Glycan
Classes in Animal Cells
OSer
OSer/Thr
NAsn
Ser-O-
OUTSIDE
INSIDE
NAsn
S S S
-O-SerS SSS S
EtnP
INOSITOL
P
NH
Ac
P
NS NS
Ac
S
2
P
GlycoproteinGlycoprotein
ProteoglycanProteoglycan
GLYCOPHOSPHO-GLYCOPHOSPHO-LIPIDLIPID
ANCHORANCHOR
N-LINKED CHAINSN-LINKED CHAINS
O-LINKED O-LINKED CHAINCHAIN
HYALURONANHYALURONAN
GLYCOSAMINO-GLYCOSAMINO-GLYCANSGLYCANS HEPARAN SULFATEHEPARAN SULFATE
CHONDROITINCHONDROITIN SULFATESULFATE
Sialic AcidsSialic Acids
GLYCOSPHINGOLIPIDGLYCOSPHINGOLIPID
O-LINKED GlcNAcO-LINKED GlcNAc
Symbolic Representation of Oligosaccharides
4β4
β 4 β2
9Ac
α3α3
α6 β 4 β2
α 6
3α
α69Acβ4 -β4
β4 β2
9Ac
α3
β4 β2 α3
α3α6α6
α3
Symbolic Representation
Fuc 3 Si3Gl4GlcNAc2Mn Fuc 6 6 Mn4GlcNAc4GlcNAc∼ 39Ac-Si6Gl4GlcNAc2Mn
Simplified Traditional
Fucp1 3 Sip2-3Glp1-4GlcNAcp1-2Mnp1 Fucp1 6 6 Mnp1-4GlcNAcp1-4GlcNAcp∼ 39OAc-Sip2-6Glp1-4GlcNAcp1-2Mnp1
Full Traditional
Clustered O-linked Oligosaccharides
• Electron micrograph of a cartilage proteoglycan shows the classical “bottle-brush” appearance
• Many mucins contain clustered O-GalNAc linked oligosaccharides
• These highly hydrated conjugates fill space, provide lubrication, and create clustered glycan ligands for binding receptors
Lecture 1 - Key Points
• What is Glycobiology?
• Central Dogma
• Basic definitions
• Monosaccharides - the structural units of glycans
• Oligoaccharides consist of monosaccharides linked together
• Nomenclature and symbolic representations
• Major classes of glycoconjugates and oligosaccharides
• Clustered oligosaccharides
• Proteins can bind at either the ends of glycans or within the chains
Outer Chains Shared by Different Classes of Glycans can be recognized by Specific
Proteins
OSer/Thr
NAsn
N-LINKED CHAINN-LINKED CHAIN
O-LINKED CHAINO-LINKED CHAIN
GLYCOSPHINGOLIPIDGLYCOSPHINGOLIPID
OUTSIDE
INSIDE
S
CELLMEMBRANE
Membrane ProteinMembrane Protein
OSer/Thr
NAsn
S
= Sialic acid
Secreted ProteinSecreted Protein
Glycosaminoglycan Chains Contain Multiple Protein Binding Sites
� Proteins can bind at the ends of N-linked and O-linked chains, and typically have shallow binding pockets
� Other proteins bind to internal sugar sequences, and have binding clefts
� Valency is a major factor in determining overall affinity
NS NSNS NSNS NSNS2S 2S2S 2S6S 6S 6S 6S 6S6S
3SAntithrombinFGF-1
Essentials of Glycobiology
March 30th, 2004
Ajit Varki
Lecture 1 Course Overview & General Introduction