Transcript Document

ESSENTIALS OF GLYCOBIOLOGY
LECTURE 33
GENETIC DISORDERS OF GLYCOSYLATION IN HUMANS
Hud Freeze
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, glycosyl
transferases, transporters, biosynthetic glycosidases
Golgi trafficking proteins.
A few disorders can be treated with monosaccharides
Misglycosylation causes some types of
muscular dystrophy
More glycosylation disorders will be found, phenotypes
Will be broad and variable
Number of defects
GLYCOSYLATION DEFECTS IDENTIFIED
N-linked Disorders
Muscular Dystrophy
20
20
15
15
10
10
5
5
80- 94
93
95
96 97 98 99 00 01 02-04
Year
CONGENITAL DISORDERS OF GLYCOSYLATION
TYPE I
LLO
Dol-PP
TYPE II
On
Protein
-N-X-T/S
-N-X-T/S
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-IIa-IIe-Ix or IIx--
>400 (Ia>300)
~20
>50
CDG--often (not always) recognized by glycosylation-dependent
alterations in serum transferrin isoelectric focusing pattern
Isoelectric focusing of transferrin
control
CDG patients
control
4
2
Sialic acids
0
normal
missing chains
Sialic acid
Galactose
altered processing
N-acetylglucosamine
Mannose
30-40 GENES REQUIRED TO MAKE AND ADD AN ASN-LINKED CHAIN
OVERVIEW OF SUGAR METABOLISM IN CELLS
Glycolysis
Gal
Glc
ATP
Man
GlcNAc
ATP
ATP
Pi
Gal-1-P
Glycogen
Glc-6-P
Man-6-P
Fru-6-P
NAD
UDP-Glc
NAD+
UDP-GlcA
GlcN-6-P
Glc-1-P
Man-l-P
GTP
AcCoA
Dol-P
ATP
GDP-Man
GlcNAc-6-P
Dol-P-Glc
NADP
-CO2
GlcNAc-l-P
GalNAc
GalNAc-1-P
UTP
GDP-Fuc
Dol-P-Man
UDP-GalNAc
UDP- GlcNAc
ManNAc
ATP
CMP-Neu5Ac
ManNAc-6-P
PEP
NADP
CMP-Neu5Ac
Dol-P
UTP
UDP-Xyl
ATP
Fuc-1-P
-NH3 Glutamine
UTP UDP-Glc
UDP-Gal
Fuc
CTP
Neu5Ac
Neu5Ac-9-P
GTP
Glycolysis
Gal
Glc
ATP
Man
GlcNAc
ATP
ATP
Pi
Gal-1-P
Glycogen
Glc-6-P
Man-6-P
Fru-6-P
Fuc-1-P
-NH3 Glutamine
UTP UDP-Glc
UDP-Gal
Fuc
NAD
UDP-Glc
NAD+
UDP-GlcA
GlcN-6-P
Glc-1-P
Man-l-P
GTP
AcCoA
Dol-P
ATP
GTP
GDP-Man
GlcNAc-6-P
Dol-P-Glc
NADP
-CO2
GlcNAc-l-P
GalNAc
GalNAc-1-P
GDP-Fuc
UTP
UDP-Xyl
ATP
Dol-P
UTP
Dol-P-Man
UDP-GalNAc
UDP- GlcNAc
ManNAc
ATP
CMP-Neu5Ac
Mannose supplements can treat
CDG-Ib, Fru-6-P-->Man-6-P defect
ManNAc-6-P
PEP
NADP
CMP-Neu5Ac
CTP
Neu5Ac
Neu5Ac-9-P
Biosynthesis and Interconve rsion of Monosaccharides. The relative contributions of
each under physiological conditions are unknown.
- monosaccharides;
-control points
and
- donors;
BEFORE MANNOSE
DURING MANNOSE
200
160
AT III
120
80
40
0
-15
-13
-11
- 13
-7
-5
-3
-1
1
3
5
1
3
5
1
3
7
9
11
13
15
17
7
anticoagulation (Marcumar6R )
5
i. v. albumin substitution
4
3
2
1
0
albumin (g/dl)
- 15
-9
- 11
-9
-7
-5
-3
-1
7
9
11
13
15
17
10
fecal alpha1 AT (mg/g)
8
6
4
2
0
-15
-13
-11
-9
-7
-5
-3
-1
time in months
5
7
9
11
13
15
17
Therapy for CDG-Ib
PMI
-6-P
Fru-6-P
PMM
-1-P
GDP-
Dol-P-
CDG defect sugar transporter
Man
GlcNAc
Glycolysis
Gal
Glc
Man
Patient
is deficient
in fucosylation
GlcNAc
ATP
ATP
What
symptoms
would Fru-6-P
you expect?
Pi
Gal-1-P
Man-6-P
Glc-6-P
Glycogen
Cause?
-NH3 Glutamine
UTP UDP-Glc
GlcN-6-P
Man-l-P
Glc-1-P
UDP-Gal
UDP-Glc
What
therapy?
NAD
NAD+
UDP-GlcA
GTP
AcCoA
Dol-P
ATP
Fuc
ATP
Fuc-1-P
GTP
GDP-Man
GlcNAc-6-P
Dol-P-Glc
NADP
-CO2
GlcNAc-l-P
GalNAc
GalNAc-1-P
UTP
GDP-Fuc
UTP
UDP-Xyl
ATP
Dol-P
Dol-P-Man
UDP-GalNAc
UDP- GlcNAc
ManNAc
ATP
Oral
fucose supplements used to
Treat
CDG-I Ic-ManNAc-6-P
PEP
Defective
in Golgi GDP-Fuc Transporter
CMP-Neu5Ac
NADP
CMP-Neu5Ac
CTP
Neu5Ac
Neu5Ac-9-P
FUCOSE THERAPY NORMALIZES NEUTROPHIL
COUNTS OF ONE CDG-IIc PATIENT
Serum Fucose
(µM)
Normal Range
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CLINICAL FEATURES OF CDG PATIENTS
Used with permission
T. Marquardt.
European J. Ped (2003)162 359-379.
Dysmorphic features of a baby with CDG-Ia (PMM-deficiency)
-6P
-6P
Ia
GDP-
Kindly provided by Dr. Thorsten Marquardt
-1P
DEFECT
-6P
-6P Ib
-1P
Ia
Dol-P-
LLO
If
Ie
GDP-
Dol-P-
Id
Dol-PP
Dol-PP
Ig
Dol-PP
Dol-PP
Dol-PP
Dol-PP
Ic
Dol-PP
Dol-PP
On
Protein
IIb
UDP
-N-X-T/S
-N-X-T/S
-N-X-T/S
-N-X-T/S
-N-X-T/S
IIc
UDP
UDP
IIa
-N-X-T/S
CMP
Cytosol
GDP-
IId
-N-X-T/S
-N-X-T/S
Golgi
GDP-
-N-X-T/S
-N-X-T/S
Defect is Shared by N-Linked and O-Linked Pathways
-N-X-T/S
T/S
-N-X-T/S
T/S
Possible Defect in Sialic Acid Synthesis or Transport?
Defects in Multiple Sialyl Transferases?
Core1GalT and S T3Gal-I activiti es in control and patient fibroblasts
(pmol/mg p rotein/min)
Enzyme
Control
H.D.
Core1GalT
3.1  0.6
1.9  0.2
ST3Gal-I
0.97  0.08
0.37  0.05
Nucleotide Sugar Transport is reduced
Endogenous acceptor
A
[3H]CMP-Sia
250
200
150
pmol/mg protein
pmol/mg protein
Exogenous acceptor (GAP)
100
50
0
250
200
B
CMP-[3H]Sia
150
100
50
0
250
C
UDP-[3H]Gal
200
D
D
300
250
200
150
100
50
0
Control
Vmax 306
HD
Vmax 92
50
40
30
20
10
0
E
200
F
CMP-Sia (uM)
Control
Vmax 43
HD Vmax 18
UDP-Gal (uM)
Control CHO
150
Vmax 182
150
100
100
50
50
0
0
0
3
Time (min)
Control
Control+TX-100
HD
6
Lec2
0
2
4
6
8
10
12
CMP-Sia (uM)
Vmax is reduced, Km remains the same
ST-GFP Trafficking is reduced in HD and FD cells
T/S
ST3Gal-I
Recovery
FD
HD
Control
Pre-bleach
bleach
10min
30min
60 min
ldlB=Cog1 ldlC=Cog2
ldlB and ldlC deficient cells are defective in glycosylation
Mutation
Function
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
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
WHEN TO CONSIDER CDG?
”EVERY TIME YOU SUSPECT IT….
AND EVERY TIME YOU DON’T”
-J. Jaeken
Glycolysis
Gal
Glc
ATP
Man
GlcNAc
ATP
ATP
Pi
Gal-1-P
Glycogen
Glc-6-P
Man-6-P
Fru-6-P
Fuc-1-P
-NH3 Glutamine
UTP UDP-Glc
UDP-Gal
Fuc
NAD
UDP-Glc
NAD+
UDP-GlcA
GlcN-6-P
Glc-1-P
Man-l-P
GTP
AcCoA
Dol-P
ATP
GTP
GDP-Man
GlcNAc-6-P
Dol-P-Glc
NADP
-CO2
GlcNAc-l-P
GalNAc
GalNAc-1-P
UTP
GDP-Fuc
UTP
UDP-Xyl
ATP
Dol-P
Dol-P-Man
UDP-GalNAc
UDP- GlcNAc
ManNAc
ATP
CMP-Neu5Ac
UDP-GlcNAc epimerase/kinase
Defective in two human diseases
ManNAc-6-P
PEP
NADP
CMP-Neu5Ac
CTP
Neu5Ac
Neu5Ac-9-P
Sialuria and Hereditary Inclusion Body Myopathy-II
Sialuria--hypotonia, cerebellar ataxia, and mental retardation
Muscular Dystrophies
Walker Warburg Syndrome--POMT1 mutations cause 30%
MEB-Muscle Eye Brain Disease--POMGnT1(1p32-34)
Encodes a b1,2GlcNAc transferase specific for O- Man
Fukuyma-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 a-dystroglycan glycosylation and not b-dystroglycan
Hereditary Inclusion Body Myopathy II (9p12-13)
UDP-GlcNAc epimerase/kinase used for CMP-Sia
O-Mannose: An Emerging Family of Glycans
WWS-Mutated transferase
O-Mannose: An Emerging Family of Glycans
MEB- Mutated transferase
MACULAR CORNEAL DYSTROPHY
Autosomal recessive
Progressive punctate corneal opacity, requires transplant
Caused by mutation in GlcNAc-6-Sulfotransferase (CHST6)
used for sulfation of keratan sulfate
http://www.burnham.org/presentations/hum
an_glycosylation_disorders/index.htm