LYSOSOMAL DEGRADATION AND PTYPE LECTINS

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LYSOSOMAL DEGRADATION AND P-TYPE LECTINS
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Trafficking of proteins
Lysosomal enzymes are secreted
and recaptured
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Normal activity on low affinity acceptor (?-MeMan)
Comparison of Cathepsin D and Glycopepsinogen to
find marker
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Why make a phosphodiester bond first then uncover
it? Why not just add P to mannose with a kinase?
Why is use a double labeled substrate donor?
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How would the number of N-linked glycans on a
lysosomal enzyme affect its affinity for one of
the M6P receptors?
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Trafficking of the Man-6-P receptors and man-6-P
proteins
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Resecretion of endocytosed ?-glucuronidase from
control and I-cell disease Fibroblasts
Why is there a lag in the re-secretion from
I-cells??
I-cell
Controls
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• USUAL TURNOVER
• Most glycans are extracellular or on cell surface
• Membrane recycling (50/hr)
• Receptor and non-receptor
• mediated endocytosis

To Endosome Lysosome
Lysosomal exoglycosidases degrade glycans at low
pH Specific lysosomal transporters carry neutral
hexose, acetylated aminohexose (GlcNAc, GalNAc),
and Anionic sugars (GlcA, Sia) to the cytoplasm.
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SUGAR CHAIN DEGRADATION ENZYMES Most
Are Lysosome/Endosome Low pH optimum,
Sugar/anomeric specificity Exo-glycosidases Targe
ted to lysosome through P-lectins and
Man-6-P But Some Are Non-lysosomal Active near
neutral pH Endoglycosidases Targeted as membrane
bound molecules Not in the lysosome
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Special Features for Degradation of Different
Glycans
TYPE FEATURE Glycoproteins N-linked ER/Golgi
/Cytoplasm/ Lysosome O-linked Unexpected
Products Proteoglycans Endoglycosidases Uniq
ue ORDER Non-glycohydrolase
enzymes Glycosphingolipids Assisting proteins
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Predict which glycans and tissues/organs are most
affected in lysosomal storage disorders affecting
multiple glycan classes
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• Special Problems for N-Linked Sugar Chains
• N-glycosylation occurs in ER-Topology for
  lysosomal degradation is wrong
• 50 of ER proteins misfold and are degraded -
  what happens to the sugar chain? To
  glycopeptides?
• Protein synthetic rate and glycosylation rate
  must be coordinated
• Competition for lectin-based chaperones

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Partially degraded glycans accumulate in tissues
and urine. Structural analysis helps to work
out the degradation pathway
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O-LINKED OLIGOSACCHARIDE DEGRADATION
Same enzymes used for N-linked oligosaccharide
degradation a-GalNAcase deficiency--produce
GalNAc terminated Oligosaccharides? Excretion
of GalNAc-a-Ser/Thr? No!! Why Not? The
oligosaccharides are larger size! How to explain
this?
a
Ser/Thr
a
Ser/Thr
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Partially degraded glycans accumulate in
tissues and urine. Structural analysis of
glycans used to work out pathway
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Degradation of Chondroitin Sulfate
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Model for the degradation of membrane-bound
GlcCer by glucocerebrosidase and SAP-C and Cer by
acid ceramidase and SAP-D, respectively. Besides
the interaction of lysosomal enzyme and activator
protein, the model emphasizes binding of
activator protein and lysosomal enzymes to the
vesicular surface containing BMP.
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What determines differences in patient outcome?
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TREATMENT FOR LYSOSOMAL STORAGE DISEASES
ENZYME REPLACMENT THERAPY
SUBSTRATE DEPLETION THERAPY
S I SE
SUBSTRATE ACCUMULATION
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ENZYME REPLACEMENT THERAPY

• WHAT PROBLEMS WOULD YOU ENCOUNTER ?
• Get it to the right cells?
• Is it stable in the cells?
• Does it need co-factors?
• Can it degrade already accumulated materials?
• Does it cross the blood brain barrier?
• Does it generate antibodies?

SUBSTRATE DEPLETION THERAPY
Potential advantages
Small molecules cross the blood-brain
barrier Targeting initial step reduces all
intermediates
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ENZYME REPLACEMENT THERAPY
FIRST ATTEMPTED IN 1980 Gaucher Disease Glucosyl
Ceramidease deficiency. Modified the N-linked
glycans targets enzyme to the liver macrophage
through a Man/GlcNAc receptor
Used in gt3000 patients as product Cerezyme
(Genzyme) Other enzymes used to treat Fabry
(a-Galactosidase), MPSI a-L-iduronidase
SUBSTRATE DEPLETION THERAPY
First Clinical trials using N-Butyldeoxynojirimyci
n, (miglustat 'Zavesca') on Gaucher patients were
successful Results for animal models for other
disorders appear promising
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Scorecard for Enzyme Replacement Therapy
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It would seem counter-intuitive to use an enzyme
inhibitor as a molecular chaperone to restore
enzyme activity in a lysosomal storage disorder. 
Explain the rationale behind this therapeutic
approach.
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EXAMPLE OF ENZYME ENHANCEMENT THERAPY
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