Transcript SIALIC ACID

Essentials of Glycobiology
Lecture 10
April 13th. 2004
Ajit Varki
The Sialic Acids
Sialic Acids
on
Vetebrate
Glycans
CHONDROITIN
SULFATE
HYALURONAN
P
GLYCOSAMINOGLYCANS
HEPARAN SULFATE
S
S
S
S
S
Ser-O-
S
S
NS
NS
-O-Ser
Proteoglycan
N-LINKED CHAINS
Ac
O-LINKED
CHAIN
GLYCOPHOSPHOLIPID
ANCHOR
P
S
O
Ser/Thr
GLYCOSPHINGOLIPID
N
Asn
N
Asn
NH 2
INOSITOL
Glycoprotein
Ac
OUTSIDE
Sialic Acids
INSIDE
O-LINKED GlcNAc
O
Ser
Etn
P
P
S
The "Primary" Sialic Acids
H
H
9
9
H-C-OH
8
H-C-OH
7
H-C-OH
H-C-OH
8
H-C-OH
7
H-C-OH
H O H
H-C-C-N
H
C5
H
6
C
O
O
C-O1
2
H
C
H3
C
OH
H
4
C
OH
a-D-Neu5Ac
6
C
O
O
HO
C5 H
4
H C
H3
C
OH
H
1
-
C-O
2
C
OH
a-D-KDN
N-acetyl-neuraminic acid
KDN
(2-keto-5-acetamido-3,5-dideoxy(2-keto-3-deoxy-D-glyceroD-glycero-D-galacto-nonulosonic acid) D-galacto-nonulosonic acid)
“Neu5Ac” “NANA”, “NeuAc”
Thought to be the metabolic
precursors of all other sialic acids
Pathways of N-acetylneuraminic Acid
(Neu5Ac) metabolism
-Neu5Ac
ENDO/PINOCYTOSIS
Neu5Ac
PLASMA MEMBRANE
SECRETORY PATHWAYS
ManNA
c
-Neu5Ac
-Neu5Ac
4
Neu5Ac
UDP-GlcNAc
LYOSOSOMES
5
Feedback Inhibition
CYTOSOL
1
ManNAc
Neu5Ac
NUCLEUS
3
CMP-Neu5Ac
2
-Neu5Ac
GOLGI
CELL
MEMBRANE
CMP-Sialic Acid Synthase
CMP-Sialic acid transporter
Sialyltransferases
Lysosomal Sialidase
5 Sialic Acid Exporter
1
2
3
4
ER
Newly synthesized glycoconjugate
Sialyltransferases in the Mammalian Genome
ST6Gal-I
ST6Gal-II
Enzyme
ST3Gal-I
ST3Gal-II
ST3Gal-III
ST3Gal-IV
ST3Gal-V
ST3Gal-VI
ST6GalNAc-I
ST6GalNAc-II
ST6GalNAc-III
ST6GalNAc-IV
ST6GalNAc-V
ST6GalNAc-VI
Galactose
N-Acetylgalactosamine
ST8Sia-I
ST8Sia-II
ST8Sia-III
ST8Sia-IV
ST8Sia-V
ST8Sia-VI
Acceptor
Galactose
Linkage
a2-6
a2-3
a2-6
Sialic acid
a2-8
Jamey Marth
Biological Roles of Sialic Acids
Structural/Physical Roles
Siglecs
Factor H
Selectins
Uterine Agglutinin
Laminins
Extrinsic
Recognition
“Non-self”
Intrinsic
Recognition
“Self”
INTRINSIC RECEPTOR
Influenza
Malaria
Cholera
Helicobacter
Mycoplasma
Rotavirus
Polyoma virus
Coronavirus
Pertussis
Tetanus etc.
EXTRINSIC RECEPTOR
SELF
M
SELF
SIALYLATED OLIGOSACCHARIDE =
M = Micro-organism
or Toxin
Molecular
Mimicry
E.Coli
Gonococcus
Meningococcus
Campylobacter
Trypanosoma
Streptococcus
Etc.
DID THE SIALIC ACIDS APPEAR LATE IN EVOLUTION?
Deuterostomes
Protostomes
Mammalia
Aves
Reptilia
Amphibia
DETECTED
Dipnoi?
Teleostei
Exceptions
Chondrostei
Elasmobranchi • Certain bacteria (mostly animal pathogens)
Agnatha
• Certain protozoa (mostly animal pathogens)
Cephalochordata
• Certain fungi (animal pathogens)
Urochordata
Hemichordata
• Rare cultured insect cell lines?
Echinodermata
• Rare insect embryo stages?
• Octopus and Squid brain?
Insecta
Crustacea
Arachinida
NOT
Annelidia
Mollusca
DETECTED
Brachiopoda
Bryozoa
Adapted from : Schauer (1982)
Microbial genes in the human genome:
lateral transfer or gene loss?
Salzberg et al. Science 2001, 292, 1903-1906
•
40 Candidate Genes fulfilling criteria suggesting
Lateral transfer from Vertebrates to Bacteria
OUR ANALYSIS OF THE LIST:
•
7 of the 40 are involved in sialic acid biosynthesis,
turnover or degradation
•
Thus, pathway involving ~0.1% of the human
genome represents almost 20% of the potential
examples of lateral gene transfer between
vertebrates and bacteria!
Phylogenetic
relationships
of enzymes
involved
in the
metabolism
of sialic acids
Neu5Ac/bacteria
Neu5Ac/Vertebrates
GlcNAc-6-phosphate
UDP-GlcNAc
GlcNAc-6phosphate
2-epimerase
UDP-GlcNAc
2-epimerase
UDP
ManNAc-6 -phosphate
ManNAc
ATP
Phosphatase
ManNAc kinase
ADP
Pi
ManNAc
ManNAc-6 -phosphate
PEP
PEP
Neu5Ac
synthetase
Pi
Pi
Neu5Ac-9phosphate
synthetase
Neu5Ac-9-phosphate
Neu5Ac-9-phosphate
phosphatase
HOMOLOGY
Pi
Neu5Ac
Angata and Varki
Chemical Reviews
102,
439-469, 2002.
CTP
Neu5Ac
CTP
CMP-Neu5Ac
synthetase
PPi
CMP-Neu5Ac
CMP-Neu5Ac
synthetase
PPi
CMP-Neu5Ac
"Universal tree" of cellular
organisms and
occurrence of sialic acids
Archae a
Biochemical
Evidence
Euryar chaeota
C renarchaeota
Genetic
Evidence
Bacteri a
Eukarya
Gram- negative
Gram- positive
Low G+C
Protozoa
Gram- positive
H ig h G+C
Protostomes
Chl amydi a
Thermus/
D einococcus
Cyanobacteri a
D eutero stomes
Aq uifex
Common ancestor
of cellular lif e
Spir ochetes
Fung i
Plants
Phylogenetic
relationships
of enzymes
involved
in the
metabolism
of sialic acids
and KDO
KDO/bacteria
Neu5Ac/bacteria
GlcNAc-6-phosphate
UDP-GlcNAc
GlcNAc-6phosphate
2-epimerase
UDP-GlcNAc
2-epimerase
UDP
ManNAc-6 -phosphate
ManNAc
ATP
Phosphatase
ManNAc kinase
ADP
Pi
ManNAc
Ara-5-phosphate
Pi
ManNAc-6 -phosphate
PEP
PEP
KDO-8-phosphate
synthetase
PEP
Neu5Ac
synthetase
Pi
Pi
Neu5Ac-9-phosphate
phosphatase
Phosphatase
Pi
Pi
Angata and Varki
Chemical Reviews
102,
439-469, 2002.
KDO
CTP
CMP-KDO
synthetase
PPi
CMP-KDO
Neu5Ac-9phosphate
synthetase
Neu5Ac-9-phosphate
KDO-8-phosphate
HOMOLOGY
Neu5Ac/Vertebrates
Neu5Ac
CTP
Neu5Ac
CTP
CMP-Neu5Ac
synthetase
PPi
CMP-Neu5Ac
CMP-Neu5Ac
synthetase
PPi
CMP-Neu5Ac
Possible Scenarios for the Phylogenetic
Origins of Sialic Acids
Scenario 1: Ancient invention
Lateral
Transfer
Amongst
Prokaryotes
Bacteria
Inventio n of Sia
Arch aea
Eu karya
Deu ter ostom e
Lin eag e
Loss or partial loss in many lineages
Biochemical
Evidence
Genetic
Evidence
Possible Scenarios for the Phylogenetic
Origins of Sialic Acids
Biochemical
Evidence
Lateral
Transfer
Amongst
Prokaryotes
Genetic
Evidence
Lateral
Transfer
to
Early
Animal
Ancestor
Loss or partial loss in many lineages
Possible Scenarios for the Phylogenetic
Origins of Sialic Acids
Biochemical
Evidence
Lateral
Transfer
Amongst
Prokaryotes
Genetic
Evidence
Lateral
Transfer
To
Prokaryotes
from
Early
Animal
Ancestor
Loss or partial loss in many lineages
Biological Roles of Sialic Acids
Structural/Physical Roles
Siglecs
Factor H
Selectins
Uterine Agglutinin
Laminins
Extrinsic
Recognition
“Non-self”
Intrinsic
Recognition
“Self”
INTRINSIC RECEPTOR
Influenza
Malaria
Cholera
Helicobacter
Mycoplasma
Rotavirus
Polyoma virus
Coronavirus
Pertussis
Tetanus etc.
EXTRINSIC RECEPTOR
SELF
M
SELF
SIALYLATED OLIGOSACCHARIDE =
M = Micro-organism
or Toxin
Molecular
Mimicry
E.Coli
Gonococcus
Meningococcus
Campylobacter
Trypanosoma
Streptococcus
Etc.
Terminal Sialic acids, Oligosialic acids, Polysialic acids
and the Enzymes that can degrade them
ENDOSIALIDASE
(ENDO-NEURAMINIDASE)
Siaa2-8Siaa2-8Siaa2-8Siaa2-8Siaa2-3(6)Galb1Ganglioside
or
N-Glycan
or
Oligosialic Acid Siaa2-8Siaa2-3(6)Galb1- O-glycan
Or
SIALIDASE (NEURAMINIDASE)
Bacterial
Glycan
Polysialic Acid
Terminal Sialic Acid Siaa2-3(6)Galb1-
The Neural
Cell Adhesion
Molecule
(N-CAM)
Expression of Polysialic
Acid on N-CAM also
affects Cell-Cell
Interactions involving
Other
Adhesion Molecules
WHY DID THE NCAM-PSA MECHANISM EVOLVE?
MOLECULAR MECHANISM FOR GLOBAL REGULATION:
PSA evolved to allow regulation of an expanded array of
different CAMs without the requirement that all of these
CAMs be affected by the same signaling pathways
EXPANDED NEED FOR PLASTICITY:
Migration of precursor cells
Pathfinding by large groups of axons
Retention of plasticity in certain adult brain tissues
Evolutionary History of N-CAM
NCAM/apCAM/FasII
Progenitor
VERTEBRATES
+PSA, +NCAM-2
INVERTEBRATES
(no PSA)
More adult PSA
(brain)
Mouse
NCAM
Human
NCAM
More “plastic”
Chimpanzee
NCAM
Aplysia
apCAM
More “hardwired”
Drosophila
FasII
Fly: “most of my genes
are like his”
Albert: “vive la différence”
Biological Roles of Sialic Acids
Structural/Physical Roles
Siglecs
Factor H
Selectins
Uterine Agglutinin
Laminins
Ligands for
Intrinsic
Receptors
Influenza
Malaria
Cholera
Helicobacter
Mycoplasma
Rotavirus
Polyoma virus
Coronavirus
Pertussis
Tetanus etc.
Ligands for
Extrinsic
Receptors
EXTRINSIC RECEPTOR
SELF
SELF
M
INTRINSIC RECEPTOR
SIALYLATED GLYCAN =
?
M = Micro-organism/Toxin
Molecular
Mimicry
E.Coli
Gonococcus
Meningococcus
Campylobacter
Trypanosoma
Streptococcus
Etc.
INFLUENZA A & B VIRUSES BIND TO CELL SURFACES VIA
A HEMAGGLUTININ THAT RECOGNIZES SIALIC ACIDS
HEMAGGLUTININ
9
INFLUENZA
A OR B
VIRUS
LINKAGE TO
UNDERLYING
SUGAR CHAIN
NEURAMINIDASE
THE "RECEPTOR-DESTROYING ENZYME"
IS A NEURAMINIDASE (SIALIDASE)
Sialic Acids Restrict Complement Activation on Cell Surfaces
FLUID
PHASE
P
C3
Bb
Factor H
Bb
C5
C3b
C3b
I
iC3b
Bb
P
Factor H
C5b
Bb
C3b C3b
Membrane
Attack
Complex
ACTIVATING
SURFACE
I
Requires
side chain Sia C3b
of Sia
NON- ACTIVATING
SURFACE
iC3b
Sialic Acid-binding Lectins are Widespread in Nature
GROUP
SUB-GROUP
Vertebrate
C - type lectins
I - type lectins
Unclassified
Adhesin
Toxin
Hemagglutinin
Hemagglutinin-esterase
Hemagglutinin-neuraminidase
Plasmodium
Tritrichomonas
Crab
Lobster
Prawn
Scorpion
Beetle
Spider
Slug
Snail
Seed
Bark
Root
Mushroom
Bacterial
Viral
Protozoal
Crustacean
Arachnid
Mollusc
Plant
Fungal
EXAMPLE
L-Selectin
CD22 (Siglec-2)
Complement Factor H
E.coli S-adhesins
Vibrio cholerae toxin
Influenza A virus
Influenza C virus
Sendai virus
P. falciparum erythrocyte-binding antigen
Tritrichomonas mobilensis Lectin.
Limulin
L-Agglutinin I
Monodin
Vaejovis spinigerus lectin
Allomyrina dichotoma lectins
Aphonopelma lectin
Limax flavus agglutinin
AchatininH
Maackia Amurensis lectin
Sambucus Nigra lectin
Tricosanthes japonicum lectin
Hericium erinaceum lectin
Natural substitutions can change the mass
and shape of the Sialic Acid molecule
N -ACETYLNEURAMINIC
ACID
9
8
7
6
2
1
5
4
3
9
8
7
6
1
2
5
4
Carbon
Oxygen
8-O-METHYL-7,9 DI-O-ACETYLN-GLYCOLYLNEURAMINIC
ACID
3
Nitrogen
Hydrogen
R7
THE SIALIC ACIDS
R9 R8 R7 R4 = hydrogen or:
R9
Acetyl
9
7
1
8
6
R9 R8 R7 R4
R5
Phosphate
2
*
5
3
R8
R2
R2
R4
OXYGEN
HYDROGEN
R8
Methyl
Sulfate
N-acetyl
R5
Hydroxyl
R8
R5
N-glycolyl
Lactyl
R9
-at physiological pH,
ionized or
lactonized or
lactamized
R1
4
R9
CARBON
R1
R5
Amino
R5
Hydrogen or:
a-linkage to:
-Gal (-3 -4 -6) or
-GalNAc (-6) or
-GlcNAc(-4 -6) or
-Sialic Acid (-8 -9)
or
b-linkage to CMP
or
Absent in:
- 2,3dehydro or
- 2,7anhydro
* (double-bond
when R2 absent)
Nomenclature and Abbreviations
Combinations of:
Neu = neuraminic acid
KDN = 2-keto-3-deoxy-nonulosonic acid.
Ac = acetyl, Gc = glycolyl, Me = methyl, Lt = lactyl, S=sulfate
Examples:
N-glycolyl-neuraminic acid = Neu5Gc
9-O-acetyl-8-O-methyl-N-acetyl-neuraminic acid = Neu5,9Ac 28Me
7,8,9-tri-O-acetyl-N-glycolyl-neuraminic acid = Neu5Gc7,8,9Ac 3
Uncertain of the type of the sialic acid ? Use generic abbreviation Sia
Sialic acid of unknown type with O- acetyl at 9-position = Sia9Ac
INFLUENZA A VIRUS
O-ACETYL ESTERS AT THE
7- AND 9-POSITIONS OF
SIALIC ACIDS
9
7
N-ACETYL-NEURAMINIC ACID
8
9-O-acetylesterases
O-acetyltransferases
9
9
8
7
7
8
INFLUENZA C
CORONAVIRUSES
O-acetyl
Migration
Migrase Enzyme?
7-O-ACETYL-N-ACETYL-NEURAMINIC ACID
9-O-ACETYL-N-ACETYL-NEURAMINIC ACID
INFLUENZA C AND CORONAVIRUS HEMAGGLUTININESTERASES SPECIFICALLY RECOGNIZE
9-O-ACETYLATED SIALIC ACIDS
HEMAGGLUTININ
9
INFLUENZA
C VIRUS
Ser
ESTERASE
LINKAGE TO
UNDERLYING
SUGAR


THE "RECEPTOR-DESTROYING ENZYME" IS A
SIALIC ACID-SPECIFIC 9-O-ACETYL-ESTERASE
O
AT 37 C, THE ESTERASE ACTIVITY IS DOMINANT
Sialic Acid-binding Proteins recognize specific
features of Sialic Acids in natural ligands: some examples
Lectins
Linkage
Underlying
Saccharide
Importance for Recognition
Carbo- N-Acyl Side
9-O-Acetylation
xylate Group Chain
of Side chain
yes
No
No
No?
Selectins
Siaa2-3
Galb1-(3)4GlcNAcb1a1
Fuc(3)4
CD22 (Siglec-2)
Siaa2-6
Galb1-4GlcNAcb1
yes
yes
yes
Blocks
Sialoadhesin (Siglec-1)
Siaa2-3
Galb1-4GlcNAcb1
yes
yes
yes
Blocks
Complement Factor H
Siaa2-?
?
yes
no?
yes
Blocks
P. falciparum
merozoite lectin
Siaa2-3
Galb1-4GlcNAcb1-
?
no?
?
Influenza A
Hemagglutinin
Siaa2-3
Siaa2-6
?
yes
variable
yes
Blocks
-
yes
no?
yes
Required
Influenza C
Hemagglutinin-esterase Siaa2-X
Blocks?
Two Major Kinds of Sialic Acids
in Mammalian Cells
N-ACETYLNEURAMINIC ACID
(Neu5Ac)
9
8
7
6
2
1
5
3
4
LINKAGE TO
UNDERLYING
SUGAR CHAIN
9
8
7
6
2
1
N-GLYCOLYLNEURAMINIC ACID
5
4
Carbon
(Neu5Gc)
3
LINKAGE TO
UNDERLYING
SUGAR CHAIN
Oxygen
Nitrogen
Hydrogen
Millions of Years Before Present*
MEAN
Amino Acid
Difference
0
5
<1.0%
Neu5Ac
Homo sapiens
Human
~0.5%
“Great Apes”
Neu5Gc
Gorilla gorilla
Pan paniscus Pan troglodytes
Bonobo
Chimpanzee
Gorilla
Pongo pygmaeus
Orangutan
Genetic
Mutation
Causing loss
Of Neu5Gc
10
*Precise Timing
Uncertain
Evolutionary
Relationships
amongst Humans
and the
Great Apes
The Sialic Acids
R1
R1
9
CARBON
7
1
8
6
R2
R1
2
5
3
NITROGEN
OXYGEN
R3
4
HYDROGEN
R1
R1 = H, ACETYL (4,7,8,9), LACTYL (9), METHYL (8), SULFATE (8,9), PHOSPHATE (9),
ANHYDRO (4,8 or 2,7), SIALIC ACID (8,9), FUCOSE(4), GLUCOSE(8), OR GALACTOSE(4)
R2 = N-ACETYL, N-GLYCOLYL, N-GLYCOLYL-O-ACETYL, AMINO, HYDROXYL
R3 = Gal (3/4/6), GalNAc(6), GlcNAc(4/6), Sia (8/9) or 5-O-Neu5Gc,
(absent in 2,6 / 2,7 ANHYDRO)