Information on EC 3.10.1.1 - N-sulfoglucosamine sulfohydrolase and Organism(s) Homo sapiens

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The expected taxonomic range for this enzyme is: Eukaryota, Bacteria


The taxonomic range for the selected organisms is: Homo sapiens

EC NUMBER
COMMENTARY hide
3.10.1.1
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RECOMMENDED NAME
GeneOntology No.
N-sulfoglucosamine sulfohydrolase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
N-sulfo-D-glucosamine + H2O = D-glucosamine + sulfate
show the reaction diagram
catalytic reaction mechanism: the active-site formylglycine (FGly70), which is intrinsically reactive, undergoes hydration to form the resting state of the enzyme with a gem-diol group (step 1). Coordination of one of the hydroxyl groups of the gem-diol to a Ca2+ ion facilitates the development of a negative charge on the O atom as its proton is lost to a base. The negatively charged O atom nucleophilically attacks the sulfur centre of the N-linked sulfate group on the glucosamine substrate (step 2), resulting in a covalently bound enzyme-substrate complex with a pentavalent sulfur transition state. An acid (possibly His181) facilitates the cleavage of the S-N bond by protonating the bridging N atom to form an amine leaving group on the N-desulfated substrate, which diffuses away, leaving an O-sulfated enzyme (step 3). Finally, in a step that underlines the importance of the formylglycine residue, another base (His125) deprotonates the second hydroxyl group, resulting in a negatively charged O atom (step 4) that forms a double bond with the C atom as the C-O bond between it and the bridging O atom of the sulfate group breaks, eliminating the sulfate ion and regenerating the formylglycine residue (step 5)
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
heparan sulfate degradation
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heparin degradation
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Glycosaminoglycan degradation
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Metabolic pathways
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SYSTEMATIC NAME
IUBMB Comments
N-sulfo-D-glucosamine sulfohydrolase
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CAS REGISTRY NUMBER
COMMENTARY hide
37289-41-1
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
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despite the low sequence identity between the unique N-sulfatase and the group of O-sulfatases, they share a similar overall fold and active-site architecture, including a catalytic formylglycine, a divalent metal-binding site and a sulfate-binding site. A highly conserved lysine in O-sulfatases is replaced in the N-sulfoglucosamine sulfohydrolase by an arginine (Arg282) that is positioned to bind the N-linked sulfate substrate
malfunction
additional information
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the enzyme shows low structural flexibility. The consensus active site lies in domain 1 in a narrow pocket at the bottom of a surface cleft and close to the end of the first beta-strand, active site structure, overview. Proposed interactions between the terminal N-sulfoglucosamine residue of the substrate with the enzyme in the active site
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
4-methylumbelliferyl-alpha-D-N-sulfoglucosaminide + H2O
4-methylumbelliferyl alpha-D-glucosaminide + sulfate
show the reaction diagram
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?
heparan sulfate + H2O
? + sulfate
show the reaction diagram
heparin + H2O
? + sulfate
show the reaction diagram
O-(alpha-2-sulfamino-2-deoxy-D-glucopyranosyl)-(1-3)-L-idonic acid
O-(alpha-2-amino-2-deoxy-D-glucopyranosyl)-(1-3)-L-idonic acid + sulfate
show the reaction diagram
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?
additional information
?
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NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
heparan sulfate + H2O
? + sulfate
show the reaction diagram
heparin + H2O
? + sulfate
show the reaction diagram
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?
additional information
?
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INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Cu2+
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1 mM decreases activity to 12% of control activity
phosphate
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SO42-
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sulfate
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.032 - 0.22
heparan sulfate
0.01
heparin
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0.0107 - 0.01225
Tetrasaccharides
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IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1
phosphate
Homo sapiens;
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pH 6.7, 37°C, recombinant enzyme
5
sulfate
Homo sapiens;
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pH 6.7, 37°C, recombinant enzyme
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4.4
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on heparin sulfate
4.9
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on heparin
5.2
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recombinant enzyme
6.7
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assay at
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37
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assay at
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
UNIPROT
ORGANISM
Homo sapiens;
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
54680
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calculation from cDNA sequence
57000
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SDS-PAGE, mature enzyme
62000
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2 * 62000, SDS-PAGE
100000
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gel filtration, fractogel TSK
110000
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gel filtration, placenta
115000
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gel filtration, Superose 12
122000
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gel filtration, Sephacryl S-300
190000
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
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x * 56000, mature recombinant enzyme, SDS-PAGE, x * 63000, unprocessed recombinant enzyme, SDS-PAGE
homodimer
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recombinant enzyme, crystal structure analysis. Two SGSH monomers associate noncovalently to form a butterfly-shaped homodimer burying approximately 10.3% of the accessible surface area of each subunit
additional information
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the monomeric enzyme subunit comprises of two domains, each centred on a beta-sheet: a large N-terminal domain (domain 1) and a smaller C-terminal domain (domain 2), as is typical for the sulfatase fold. There are 14 beta-strands, 13 alpha-helices and six 310-helices (T1-T6) in total
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycoprotein
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified recombinant C-terminally His6-tagged, glycosylated full-length enzyme, small unit and large unit crystal forms, X-ray diffraction structure determination and analysis at 2.0-2.4 A resolution, molecular replacement. The enzyme appears to exist as a homodimer in both crystal forms
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
4, sodium acetate, pH 5.0, 30% glycerol, 25 mM NaCl, 0.1 mM DTT, 0.3 mg/ml BSA, 3 month
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
apparent homogeneity
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purification of recombinant enzyme
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recombinant C-terminally His6-tagged full-length enzyme from HEK-293 cells by nickel affinity croomatography
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SP-Sepharose column chromatography and Sephacryl S-300 gel filtration
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
DNA and amino acid sequence determination and anaylsis, genotyping of patients with mucopolysaccharidosis type IIIA, overview
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expressed in CHO-K1 cells
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expression in Chinese hamster ovary cells
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expression of wild-type and mutant enzymes in BHK or CHO cells
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gene SGSH, DNA and amino acid sequence determination and analysis, expression of mutant S298P sulfamidase in BHK cells, rapid degradation is responsible for the low steady state level of the mutant protein. Only small amounts of the S298P sulfamidase are transported to the lysosomes after processing and secretion, most of the mutant sulfamidase exits the endoplasmic reticulum for proteasomal degradation
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heterologous expression, using a baculovirus system, of the cDNAs corresponding to eight out of 14 mutant alleles present in the patient group of 26 unrelated Spanish patients with mucopolysaccharidosis IIIA. Expression of the following alleles: p.S66W, p.R74H, p.Q85R, p.R206P, p.L386R, p.R433W, p.R433Q, and c.1079delC. Expression of wild-type and mutant sulfamidases in Sf9 insect cells
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recombinant expression of C-terminally His6-tagged full-length enzyme in HEK-293 cells
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
G191R
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mutation identified in children with mucopolysaccharidosis type IIIA in Germany. Recombinant mutant enzyme shows no activity
L163P
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mutation identified in children with mucopolysaccharidosis type IIIA in Germany. Recombinant mutant enzyme shows no activity
R206P
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an adult Sanfilippo type A patient with homozygous mutation R206P in the sulfamidase gene shows a mild clinical phenotype characterized essentially by a moderate nonevolving metal retardation, polymorphism R456H is also found
R233X
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mutation identified in children with mucopolysaccharidosis type IIIA in Germany. Recombinant mutant enzyme shows no activity
R433W
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mutation identified in children with mucopolysaccharidosis type IIIA in Germany. Recombinant mutant enzyme shows no activity
R74C
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mutation identified in children with mucopolysaccharidosis type IIIA in Germany. Recombinant mutant enzyme shows no activity
S106R
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mutation identified in children with mucopolysaccharidosis type IIIA in Germany. Recombinant mutant enzyme shows 3.3% of wild-type activity
Y403del
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mutation identified in children with mucopolysaccharidosis type IIIA in Germany. Recombinant mutant enzyme shows no activity
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
diagnostics
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a fluorimetric SGSH activity assay is commonly used to examine cells of mucopolysaccharidosis type IIIA patients for N-sulfoglucosamine sulfohydrolase activity, modification of the method for brain homogenates and definition of the parameters for assay linearity, overview
drug development
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the enzyme is a target for the development of structure-based drug design for the devastating neurodegenerative disorder mucopolysaccharidosis type IIIA or Sanfilippo A syndrome
medicine