essentials of glycobiology lecture 33 genetic disorders of glycosylation in humans hud freeze
TRANSCRIPT
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ESSENTIALS OF GLYCOBIOLOGY
LECTURE 33
GENETIC DISORDERS OF GLYCOSYLATION IN HUMANS
Hud Freeze
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OVERVIEW AND SUMMARY
Many Human glycosylation disorders were discovered in the last 10 years, mostly in the N-linked pathway.
Disorders are underdiagnosed. Alert physicians.
Defects include monosaccharide activation, glycosyltransferases, transporters, biosynthetic glycosidasesGolgi trafficking proteins.
A few disorders can be treated with monosaccharides
Misglycosylation causes some types of muscular dystrophy
More glycosylation disorders will be found, phenotypesWill be broad and variable
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GLYCOSYLATION DEFECTS IDENTIFIED GLYCOSYLATION DEFECTS IDENTIFIED
Year99989796959480-
930100
15
5
10
20
5
10
15
20N-linked DisordersMuscular Dystrophy
02-04
Nu
mb
er o
f d
efec
ts
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-N-X-T/S
On Protein
Dol-P-P
LLO
TYPE I TYPE II
CONGENITAL DISORDERS OF GLYCOSYLATION
-N-X-T/S
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CONGENITAL DISORDERS OF GLYCOSYLATION(CDG)
TYPE I---Defects in synthesis or transfer of oligosaccharides from Dol-PP carrier to proteins in the lumen of the endoplasmic reticulum
TYPE II--Defects in the processing of N-linked oligosaccharides OR in all other types of glycosylation
Types Number of Patients
Ia-IL-- >400 (Ia>300) IIa-IIe-- ~20 Ix or IIx-- >50
CDG--often (not always) recognized by glycosylation-dependentalterations in serum transferrin isoelectric focusing pattern
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Isoelectric focusing of transferrinIsoelectric focusing of transferrin
4
2
0
control controlCDG patients
Sialic acidGalactose
N-acetylglucosamineMannose
normal missing chains altered processing
Sialic acids
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30-40 GENES REQUIRED TO MAKE AND ADD AN ASN-LINKED CHAIN30-40 GENES REQUIRED TO MAKE AND ADD AN ASN-LINKED CHAIN30-40 GENES REQUIRED TO MAKE AND ADD AN ASN-LINKED CHAIN30-40 GENES REQUIRED TO MAKE AND ADD AN ASN-LINKED CHAIN
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Man Fuc
GDP-Man
Man-l-P
GTP
NADP
Dol-P-Man
GDP-Fuc
Man-6-P
Dol-P
ATP
Fuc-1-P
GTP
ATP
ManNAc-6-P
PEP
Neu5AcCMP-Neu5Ac Neu5Ac-9-P
CTP
ManNAc
UDP- GlcNAcUDP-GalNAcGalNAc
ATP
GalNAc-1-P
UTP
GlcNAc-6-P
GlcNAc-l-P
UTP
Dol-P-Glc
GlcNAc
UDP-Xyl
-CO2
UDP-GlcA
UDP-Glc
Glycogen
NAD+
UDP-Gal
Gal
Gal-1-P
UTP UDP-Glc
Dol-P
ATP
Glc-1-P
Glc
Glc-6-P Fru-6-P
ATP
Glutamine
GlcN-6-P
-NH3
Pi
ATP
AcCoA
NAD
Glycolysis
CMP-Neu5Ac
NADP
OVERVIEW OF SUGAR METABOLISM IN CELLS
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Man Fuc
GDP-Man
Man-l-P
GTP
NADP
Dol-P-Man
GDP-Fuc
Man-6-P
Dol-P
ATP
Fuc-1-P
GTP
ATP
ManNAc-6-P
PEP
Neu5AcCMP-Neu5Ac Neu5Ac-9-P
CTP
ManNAc
UDP- GlcNAcUDP-GalNAcGalNAc
ATP
GalNAc-1-P
UTP
GlcNAc-6-P
GlcNAc-l-P
UTP
Dol-P-Glc
GlcNAc
UDP-Xyl
-CO2
UDP-GlcA
UDP-Glc
Glycogen
NAD+
UDP-Gal
Gal
Gal-1-P
UTP UDP-Glc
Dol-P
ATP
Glc-1-P
Glc
Glc-6-P Fru-6-P
ATP
Glutamine
GlcN-6-P
-NH3
Pi
ATP
AcCoA
NAD
Glycolysis
CMP-Neu5Ac
NADP
Biosynthesis and Interconversion of Monosaccharides. The relative contributions of
each under physiological conditions are unknown. and - donors;
- monosaccharides; -control points
Mannose supplements can treat CDG-Ib, Fru-6-P-->Man-6-P defect
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0
40
80
120
160
200
-15 -13 -11 -9 -7 -5 -3 -1 1 3 5 7 9 11 13 15 17
0
1
2
3
4
5
6
7
-15 -13 -11 -9 -7 -5 -3 -1 1 3 5 7 9 11 13 15 17
0
2
4
6
8
10
-15 -13 -11 -9 -7 -5 -3 -1 1 3 5 7 9 11 13 15 17
anticoagulation (Marcumar R )
i. v. albumin substitution
albumin (g/dl)
fecal alpha1 AT (mg/g)
AT III
time in months
BEFORE MANNOSE DURING MANNOSE
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CDG defect sugar transporter Man GlcNAc
-6-P
-1-P GDP-
Dol-P-
Fru-6-PPMI
PMM
Therapy for CDG-Ib
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Man Fuc
GDP-Man
Man-l-P
GTP
NADP
Dol-P-Man
GDP-Fuc
Man-6-P
Dol-P
ATP
Fuc-1-P
GTP
ATP
ManNAc-6-P
PEP
Neu5AcCMP-Neu5Ac Neu5Ac-9-P
CTP
ManNAc
UDP- GlcNAcUDP-GalNAcGalNAc
ATP
GalNAc-1-P
UTP
GlcNAc-6-P
GlcNAc-l-P
UTP
Dol-P-Glc
GlcNAc
UDP-Xyl
-CO2
UDP-GlcA
UDP-Glc
Glycogen
NAD+
UDP-Gal
Gal
Gal-1-P
UTP UDP-Glc
Dol-P
ATP
Glc-1-P
Glc
Glc-6-P Fru-6-P
ATP
Glutamine
GlcN-6-P
-NH3
Pi
ATP
AcCoA
NAD
Glycolysis
CMP-Neu5Ac
NADP
Oral fucose supplements used to Treat CDG-IIc-- Defective in Golgi GDP-Fuc Transporter
Patient is deficient in fucosylationWhat symptoms would you expect?Cause?What therapy?
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FUCOSE THERAPY NORMALIZES NEUTROPHILCOUNTS OF ONE CDG-IIc PATIENT
Serum Fucose(µM)
Normal Range
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QuickTime™ and aBMP decompressor
are needed to see this picture.
QuickTime™ and aPhoto - JPEG decompressor
are needed to see this picture.
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CLINICAL FEATURES OF CDG PATIENTS
Used with permission T. Marquardt.
European J. Ped (2003)162 359-379.
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Kindly provided by Dr. Thorsten Marquardt
Dysmorphic features of a baby with CDG-Ia (PMM-deficiency)
-6P
GDP-
Ia-6P -1P
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DEFECT
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-6P -1P
GDP-
GDP-
-N-X-T/S
Golgi
CytosolIIa
IIb
IIcUDP
CMPUDP
UDP
-N-X-T/S -N-X-T/S
-N-X-T/S -N-X-T/S
-N-X-T/S -N-X-T/S
-N-X-T/S -N-X-T/S
-N-X-T/S
IId
Dol-P-P
-6P Dol-P-
GDP-
Dol-P-Ib Ia
Ic
Id
Ie If
Dol-P-P
IgDol-P-P
Dol-P-P
Dol-P-P
Dol-P-P
Dol-P-P
Dol-P-P
LLO
On Protein
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-N-X-T/S -N-X-T/S
T/ST/S
Possible Defect in Sialic Acid Synthesis or Transport?
Defect is Shared by N-Linked and O-Linked Pathways
Defects in Multiple Sialyl Transferases?
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Core1GalT and ST3Gal-I activities in control and patient fibroblasts(pmol/mg protein/min)
Enzyme Control H.D.
Core1GalT 3.1 ± 0.6 1.9 ± 0.2
ST3 -Gal I 0.97 ± 0.08 0.37 ± 0.05
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pm
ol/
mg
pro
tein
Endogenous acceptor
pm
ol/
mg
pro
tein
0 4 6 8 10 122
Exogenous acceptor (GAP)
D
Vmax 182
Lec20
50
100
150
200Control CHO
D
E
F
ControlVmax 43
Control
Vmax 306
HD Vmax 18
HDVmax 92
0
50
150
250300
200
100
01020
304050
CMP-Sia (uM)
UDP-Gal (uM)
CMP-Sia (uM)
60Time (min)
0
50
100
150
200
250
CMP-[3H]Sia
0
50
100
150
200
250[3H]CMP-Sia
0
50
100
150
200
250
UDP-[3H]Gal
A
B
C
3
Control Control+TX-100 HD
Nucleotide Sugar Transport is reduced
Vmax is reduced, Km remains the same
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H
DC
ontr
ol
F
D Pre-bleach bleach 10min 30min 60 min
Recovery
ST-GFP Trafficking is reduced in HD and FD cells
T/SST3Gal-IST3Gal-I
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ldlB=Cog1 ldlC=Cog2
ldlB and ldlC deficient cells are defective in glycosylation
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Cog7 mutation decreases mRNA and protein
Destabilizes COG complex, mislocalizing some COG subunits
Decreases trafficking of glycosyltransferases and transporters
Decreases enzyme/transporter stability and normal localization
Decreases terminal modifications on glycans
Mutation Function
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SUMMARY• Physicians must know many faces of glycosylation • Defective Glycosylation causes disease • Only a few of the potential types of CDG are known• Simple therapy may help a few CDG patients• CDG may be the cause of unexplained cases of developmental
delay, coagulopathy, and hypoglycemia• CDG presentations can be very mild and broad
WHENWHEN TO CONSIDER CDG?TO CONSIDER CDG?
””EVERY TIME YOU SUSPECT IT….EVERY TIME YOU SUSPECT IT….AND EVERY TIME YOU DON’T”AND EVERY TIME YOU DON’T”
-J. Jaeken-J. Jaeken
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Man Fuc
GDP-Man
Man-l-P
GTP
NADP
Dol-P-Man
GDP-Fuc
Man-6-P
Dol-P
ATP
Fuc-1-P
GTP
ATP
ManNAc-6-P
PEP
Neu5AcCMP-Neu5Ac Neu5Ac-9-P
CTP
ManNAc
UDP- GlcNAcUDP-GalNAcGalNAc
ATP
GalNAc-1-P
UTP
GlcNAc-6-P
GlcNAc-l-P
UTP
Dol-P-Glc
GlcNAc
UDP-Xyl
-CO2
UDP-GlcA
UDP-Glc
Glycogen
NAD+
UDP-Gal
Gal
Gal-1-P
UTP UDP-Glc
Dol-P
ATP
Glc-1-P
Glc
Glc-6-P Fru-6-P
ATP
Glutamine
GlcN-6-P
-NH3
Pi
ATP
AcCoA
NAD
Glycolysis
CMP-Neu5Ac
NADP
UDP-GlcNAc epimerase/kinase Defective in two human diseases
Sialuria and Hereditary Inclusion Body Myopathy-II
Sialuria--hypotonia, cerebellar ataxia, and mental retardation
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Muscular Dystrophies
Walker Warburg Syndrome--POMT1 mutations cause 30%MEB-Muscle Eye Brain Disease--POMGnT1(1p32-34)
Encodes a 1,2GlcNAc transferase specific for O-ManFukuyma-type CMD--fukutin (9q31)
Putative transferase in golgi (?)Fukutin Related protein--(19q13.3)
Putative transferase (?)LARGE- cause of myd mouse, tandem glycosyltransfrases(?)
Common Features:Affects -dystroglycan glycosylation and not -dystroglycan
Hereditary Inclusion Body Myopathy II (9p12-13)UDP-GlcNAc epimerase/kinase used for CMP-Sia
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O-Mannose: An Emerging Family of Glycans
WWS-Mutated transferase
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O-Mannose: An Emerging Family of Glycans
MEB- Mutated transferase
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MACULAR CORNEAL DYSTROPHY
Autosomal recessiveProgressive punctate corneal opacity, requires transplantCaused by mutation in GlcNAc-6-Sulfotransferase (CHST6)
used for sulfation of keratan sulfate
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http://www.burnham.org/presentations/human_glycosylation_disorders/index.htm