Information on EC 2.4.1.175 - glucuronosyl-N-acetylgalactosaminyl-proteoglycan 4-beta-N-acetylgalactosaminyltransferase

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

EC NUMBER
COMMENTARY hide
2.4.1.175
-
RECOMMENDED NAME
GeneOntology No.
glucuronosyl-N-acetylgalactosaminyl-proteoglycan 4-beta-N-acetylgalactosaminyltransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
UDP-N-acetyl-alpha-D-galactosamine + beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-galactosaminyl-proteoglycan = UDP + N-acetyl-beta-D-galactosaminyl-(1->4)-beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-galactosaminyl-proteoglycan
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hexosyl group transfer
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
chondroitin biosynthesis
-
-
Glycosaminoglycan biosynthesis - chondroitin sulfate / dermatan sulfate
-
-
Metabolic pathways
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-
SYSTEMATIC NAME
IUBMB Comments
UDP-N-acetyl-D-galactosamine:beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-galactosaminyl-proteoglycan 4-beta-N-acetylgalactosaminyltransferase
Involved in the biosynthesis of chondroitin sulfate. The human form of this enzyme is a bifunctional glycosyltransferase, which also has the 3-beta-glucuronosyltransferase (EC 2.4.1.226, N-acetylgalactosaminyl-proteoglycan 3-beta-glucuronosyltransferase) activity required for the synthesis of the chondroitin sulfate disaccharide repeats. Similar chondroitin synthase 'co-polymerases' can be found in Pasteurella multocida and Escherichia coli.
CAS REGISTRY NUMBER
COMMENTARY hide
96189-40-1
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
type F
SwissProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
-
the enzyme is a member of the ChSy family
malfunction
metabolism
-
CSS1 and CSS2 differently contribute to chondroitin sulfate biosynthesis, mechanism of chondroitin sulfate biosynthesis, overview. CSS1 participates in both the chondroitin sulfate chain initiation and polymerization, and CSS2 participates in the polymerization, especially in further extension
physiological function
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
alpha-thrombomodulin + UDP-N-acetyl-D-galactosamine + UDP-glucuronic acid
?
show the reaction diagram
-
-
-
?
chondroitin + UDP-GalNAc
?
show the reaction diagram
chondroitin is the best substrate
-
-
?
chondroitin + UDP-glucuronic acid
?
show the reaction diagram
chondroitin is the best substrate
-
-
?
chondroitin sulfate A + UDP-GalNAc
?
show the reaction diagram
-
-
-
?
chondroitin sulfate A + UDP-glucuronic acid
?
show the reaction diagram
-
-
-
?
chondroitin sulfate A10 + UDP-GalNAc
?
show the reaction diagram
no activity with UDP-glucuronic acid
-
-
?
chondroitin sulfate A11 + UDP-glucuronic acid
?
show the reaction diagram
no activity with UDP-GalNAc
-
-
?
chondroitin sulfate C + UDP-GalNAc
?
show the reaction diagram
-
-
-
?
chondroitin sulfate C + UDP-glucuronic acid
?
show the reaction diagram
-
-
-
?
chondroitin sulfate C10 + UDP-GalNAc
?
show the reaction diagram
no activity with UDP-glucuronic acid
-
-
?
chondroitin sulfate C11 + UDP-glucuronic acid
?
show the reaction diagram
no activity with UDP-GalNAc
-
-
?
chondroitin sulfate D + UDP-GalNAc
?
show the reaction diagram
-
-
-
?
chondroitin sulfate D + UDP-glucuronic acid
?
show the reaction diagram
-
-
-
?
chondroitin sulfate E + UDP-GalNAc
?
show the reaction diagram
-
-
-
?
chondroitin sulfate E + UDP-glucuronic acid
?
show the reaction diagram
-
-
-
?
dermatan sulfate + UDP-GalNAc
?
show the reaction diagram
-
-
-
?
dermatan sulfate + UDP-glucuronic acid
?
show the reaction diagram
-
-
-
?
GlcUAbeta1-3Galbeta1-O-C2H4NH-benzyloxycarbonyl + UDP-glucuronic acid
?
show the reaction diagram
-
-
-
-
?
GlcUAbeta1-3Galbeta1-O-C2H4NH-benzyloxycarbonyl + UDP-N-acetyl-D-galactosamine
?
show the reaction diagram
-
-
-
-
?
GlcUAbeta1-3Galbeta1-O-naphthalenemethanol + UDP-glucuronic acid
?
show the reaction diagram
-
-
-
-
?
GlcUAbeta1-3Galbeta1-O-naphthalenemethanol + UDP-N-acetyl-D-galactosamine
?
show the reaction diagram
-
-
-
-
?
hyaluronan + UDP-GalNAc
?
show the reaction diagram
-
-
-
?
hyaluronan + UDP-glucuronic acid
?
show the reaction diagram
-
-
-
?
UDP-alpha-D-glucuronate + chondroitin hexasaccharide
UDP + ?
show the reaction diagram
UDP-alpha-D-glucuronate + N-acetyl-beta-D-galactosaminyl-[GlcA-GalNAc]n-pyridylamine
UDP + [GlcA-GalNAc]n+1-pyridylamine
show the reaction diagram
-
-
-
-
?
UDP-N-acetyl-alpha-D-galactosamine + (glucuronyl-N-acetyl-1,3-beta-D-galactosamine 4-sulfate)4
UDP + N-acetyl-beta-D-galactosaminyl-1,4-glucuronyl-N-acetyl-1,3-beta-D-galactosaminyl-(glucuronyl-N-acetyl-1,3-beta-D-galactosamine)3
show the reaction diagram
-
i.e. chondroitin sulfate octasaccharide, glycosylated at 32% the rate of chondroitin octasaccharide
-
-
?
UDP-N-acetyl-alpha-D-galactosamine + (glucuronyl-N-acetyl-1,3-beta-D-galactosamine)2
UDP + N-acetyl-beta-D-galactosaminyl-1,4-glucuronyl-N-acetyl-1,3-beta-D-galactosaminyl-glucuronyl-N-acetyl-1,3-beta-D-galactosamine
show the reaction diagram
UDP-N-acetyl-alpha-D-galactosamine + (glucuronyl-N-acetyl-1,3-beta-D-galactosamine)3
UDP + N-acetyl-beta-D-galactosaminyl-1,4-glucuronyl-N-acetyl-1,3-beta-D-galactosaminyl-(glucuronyl-N-acetyl-1,3-beta-D-galactosamine)2
show the reaction diagram
UDP-N-acetyl-alpha-D-galactosamine + (glucuronyl-N-acetyl-1,3-beta-D-galactosamine)4
UDP + N-acetyl-beta-D-galactosaminyl-1,4-glucuronyl-N-acetyl-1,3-beta-D-galactosaminyl-(glucuronyl-N-acetyl-1,3-beta-D-galactosamine)3
show the reaction diagram
UDP-N-acetyl-alpha-D-galactosamine + (glucuronyl-N-acetyl-1,3-beta-D-galactosamine)5
UDP + N-acetyl-beta-D-galactosaminyl-1,4-glucuronyl-N-acetyl-1,3-beta-D-galactosaminyl-(glucuronyl-N-acetyl-1,3-beta-D-galactosamine)4
show the reaction diagram
UDP-N-acetyl-alpha-D-galactosamine + (glucuronyl-N-acetyl-1,3-beta-D-galactosamine)6
UDP + N-acetyl-beta-D-galactosaminyl-1,4-glucuronyl-N-acetyl-1,3-beta-D-galactosaminyl-(glucuronyl-N-acetyl-1,3-beta-D-galactosamine)5
show the reaction diagram
UDP-N-acetyl-alpha-D-galactosamine + (glucuronyl-N-acetyl-1,3-beta-D-galactosamine)7
UDP + N-acetyl-beta-D-galactosaminyl-1,4-glucuronyl-N-acetyl-1,3-beta-D-galactosaminyl-(glucuronyl-N-acetyl-1,3-beta-D-galactosamine)6
show the reaction diagram
UDP-N-acetyl-alpha-D-galactosamine + (glucuronyl-N-acetyl-1,3-beta-D-galactosamine)8
UDP + N-acetyl-beta-D-galactosaminyl-1,4-glucuronyl-N-acetyl-1,3-beta-D-galactosaminyl-(glucuronyl-N-acetyl-1,3-beta-D-galactosamine)7
show the reaction diagram
-
i.e. chondroitin hexadecasaccharide, glycosylated at 15% the rate of decamer glycosylation
-
-
?
UDP-N-acetyl-alpha-D-galactosamine + (glucuronyl-N-acetyl-1,3-beta-D-glucosamine)4
UDP + ?
show the reaction diagram
-
i.e. hyaluronate octasaccharide, glycosylated at 25% the rate of chondroitin octasaccharide
-
-
?
UDP-N-acetyl-alpha-D-galactosamine + beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-galactosaminyl-proteoglycan
UDP + N-acetyl-beta-D-galactosaminyl-(1->4)-beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-galactosaminyl-proteoglycan
show the reaction diagram
UDP-N-acetyl-alpha-D-galactosamine + chondroitin hexasaccharide
UDP + N-acetyl-beta-D-galactosaminyl chondroitin hexasaccharide
show the reaction diagram
UDP-N-acetyl-alpha-D-galactosamine + chondroitin oligosaccharide
UDP + N-acetyl-beta-D-galactosaminyl chondroitin oligosaccharide
show the reaction diagram
UDP-N-acetyl-alpha-D-galactosamine + chondroitin pentasaccharide
UDP + N-acetyl-beta-D-galactosaminyl chondroitin pentasaccharide
show the reaction diagram
UDP-N-acetyl-alpha-D-galactosamine + chondroitin polysaccharide
UDP + N-acetyl-beta-D-galactosaminyl chondroitin polysaccharide
show the reaction diagram
UDP-N-acetyl-alpha-D-galactosamine + chondroitin sulfate oligosaccharide
UDP + N-acetyl-beta-D-galactosaminyl chondroitin sulfate oligosaccharide
show the reaction diagram
UDP-N-acetyl-alpha-D-galactosamine + [GlcA-GalNAc]n-pyridylamine
UDP + N-acetyl-beta-D-galactosaminyl-[GlcA-GalNAc]n-pyridylamine
show the reaction diagram
UDP-N-acetyl-alpha-D-glucosamine + chondroitin sulfate oligosaccharide
UDP + N-acetyl-beta-D-glucosaminyl chondroitin sulfate oligosaccharide
show the reaction diagram
UDP-N-acetyl-D-galactosamine + beta-D-glucuronyl-(1->3)-N-acetyl-beta-D-galactosaminyl-proteoglycan
UDP + N-acetyl-D-galactosaminyl-(1->4)-beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-galactosaminyl-proteoglycan
show the reaction diagram
-
-
-
-
?
UDP-N-acetyl-D-galactosamine + chondroitin oligosaccharide
UDP + N-acetyl-D-galactosaminyl chondroitin oligosaccharide
show the reaction diagram
UDP-N-acetyl-D-galactosamine + chondroitin polysaccharide
UDP + N-acetyl-D-galactosaminyl chondroitin polysaccharide
show the reaction diagram
best acceptor substrate
-
?
UDP-N-acetyl-D-galactosamine + chondroitin sulfate C
UDP + N-acetyl-D-galactosaminyl chondroitin sulfate C
show the reaction diagram
UDP-N-acetyl-D-galactosamine + chondroitin sulfate oligosaccharide
UDP + N-acetyl-D-galactosaminyl chondroitin sulfate oligosaccharide
show the reaction diagram
UDP-N-acetyl-D-galactosamine + defructosylated K4 oligosaccharide
UDP + N-acetyl-D-galactosaminyl defructosylated K4 oligosaccharide
show the reaction diagram
additional information
?
-
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
UDP-N-acetyl-alpha-D-galactosamine + beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-galactosaminyl-proteoglycan
UDP + N-acetyl-beta-D-galactosaminyl-(1->4)-beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-galactosaminyl-proteoglycan
show the reaction diagram
UDP-N-acetyl-alpha-D-galactosamine + chondroitin oligosaccharide
UDP + N-acetyl-beta-D-galactosaminyl chondroitin oligosaccharide
show the reaction diagram
UDP-N-acetyl-alpha-D-galactosamine + chondroitin sulfate oligosaccharide
UDP + N-acetyl-beta-D-galactosaminyl chondroitin sulfate oligosaccharide
show the reaction diagram
UDP-N-acetyl-D-galactosamine + beta-D-glucuronyl-(1->3)-N-acetyl-beta-D-galactosaminyl-proteoglycan
UDP + N-acetyl-D-galactosaminyl-(1->4)-beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-galactosaminyl-proteoglycan
show the reaction diagram
-
-
-
-
?
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Cd2+
-
activates
Co2+
-
divalent cation is essential, Co2+ evokes the highest level of activity
Fe2+
-
can partly substitute for Mn2+, 30.6% activity compared to Mn2+
Mg2+
-
can partly substitute for Mn2+, 30.7% activity compared to Mn2+
additional information
-
Mg2+, Cu2+, Ca2+, Zn2+, Ba2+ do not effect the enzyme activity
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Ca2+
-
complete inhibition
Cu2+
-
complete inhibition
EDTA
-
complete inhibition
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
chondroitin polymerizing factor (ChPF) is required for chondroitin polymerizing activity of mammalian ChSy. Chondroitin polymerization is not demonstrated in vitro using the recombinant enzyme
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.36
chondroitin sulfate CS-11
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-
-
0.221
UDP-GalNAc
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-
0.005895 - 0.05
UDP-N-acetyl-beta-D-galactosamine
0.005895
UDP-N-acetyl-beta-D-glucosamine
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wild type enzyme, in 25 mM HEPES (pH 7.5), 100 mM NaCl, 20 mM MnCl2, 1 mM CaCl2, at 30C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.000006
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recombinant CSS2B variant in COS-7 cells, pH 6.2, 37C
0.000008
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recombinant CSS2A variant in COS-7 cells, pH 6.2, 37C
0.00053
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purified rcombinant enzyme, substrate chondroitin polysaccharide with MW 10 kDa
0.00124
-
purified rcombinant enzyme, substrate chondroitin hexasaccharide
0.00331
-
purified rcombinant enzyme, substrate chondroitin sulfate polysaccharide with MW 20 kDa
0.00334
-
purified rcombinant enzyme, substrate chondroitin sulfate hexasaccharide
0.0425
-
purified enzyme
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.8
-
Mes buffer, reaction with chondroitin sulfate C11 and UDP-glucuronic acid
7 - 7.5
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-
7.4
-
-
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25
-
highest product chain length
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
melanoma cell line
Manually annotated by BRENDA team
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soft tissue sarcomas with myxoid substance: myxoid liposarcoma, 14 of myxofibrosarcoma, 12 of malignant peripheral nerve sheath tumor, and 2 of low-grade fibromyxoid sarcoma. Soft tissue sarcoma is a malignant tumor of nonepithelial extraskeletal origin classified by specific, differentiated cell types
Manually annotated by BRENDA team
-
-
Manually annotated by BRENDA team
thyroid cancer cell line
Manually annotated by BRENDA team
myeloma cell line
Manually annotated by BRENDA team
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
-
-
-
Manually annotated by BRENDA team
additional information
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
82000
-
SDS-PAGE
160000
-
gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycoprotein
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
K4CP in the presence of UDP and UDP-N-acetylgalactosamine as well as with UDP and UDP-glucuronic acid, hanging drop vapor diffusion method and batch method, using 15-20% (w/v) PEG 3350 and 200 mM NaCl
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50
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t1/2: 5 min, 90% loss of activity within 20 min
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
ion-exchange chromatography on DEAE-trisacryl inactivates
-
rapid thermodenaturation
-
stable against freeze-thawing in liquid N2 several times
-
unstable upon purification
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-80C, MES/NaOH, 50 mM, pH 6.5, 1 mM dithiothreitol, stable for at least a few months
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
glutathione Sepharose column chromatography and Superdex 200 gel filtration
-
glutathione-Sepharose resin column chromatography
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Ni-NTA agarose column chromatography
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partially solubilized with a buffer containing 2% Triton X-100, 20% glycerol and phospholipase A2
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recombinant His-tagged enzyme expressed in E. coli strain TOP10
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recombinant protein using protein A-tag
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ChGn-2, expression analysis by quantitative real-time RT-PCR, co-expression of ChGn-1 or ChGn-2 into sog9 and L cells, sog9 is a mutant L cell line are deficient in the expression of chondroitin 4-O-sulfotransferase-1, C4ST-1, disaccharide composition of chondroitin sulfate in control and transfected L cells, overview
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CSS2 quantitative expression analysis by realtime RT-PCR, overview
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DNA and amino acid sequence determination and analysis, expression as soluble His-tagged protein in Escherichia coli strain TOP10
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DNA sequence determination and analysis, expression of a truncated form, consisting of the putative catalytic domain, in Spodoptera frugiperda Sf21 insect cells as a soluble FLAG-tagged protein
DNA sequence determination, expression in Escherichia coli of amino acid residues 1-704 of total 965 as soluble functional protein
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DNA sequence determination, genomic organization and chromosome localization, functional expression as soluble protein fused to protein A IgG-binding domain in COS-1 cells, secretion into the medium
expressed in Escherichia coli BL21(DE3) cells
-
expressed in Escherichia Top10 cells
-
expressed in HeLa cells; soluble form of the enzyme lacking the first 129 amino acids expressed as protein A fusion protein in COS-1 cells
expression in COS cells
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gene ChSy-1, genotyping in healthy persons and multiple sclerosis patients, recombinant expression of wild-type and mutant enzymes in COS-1 cells
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gene CHSY1, quantitative reverse-transcription PCR enzyme expression analysis
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gene chsy1, real-time PCR enzyme expression analysis
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gene CHSY1, soluble forms of wild-type and mutant ChSy-1 is heterologously expressed in COS-1 cells, the recombinant enzymes are secreted to the medium, quantitative real-time RT-PCR enzyme expression analysis
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genomic organization and chromosome localization, functional expression as soluble protein fused to protein A IgG-binding domain in COS-1 cells, secretion into the medium
GFP-tagged and DsRed-monomer tagged enzyme expressed in HeLa cells; soluble form of the enzyme lacking the first 57 amino acids expressed as protein A fusion protein in COS-1 cells
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individual overexpression of soluble forms of FLAG- and Myc-tagged splice variants CSS2A and CSS2B in COS-7 cells, co-expression with CSS1, EC 2.4.1.174. Whereas CSS2A facilitates chondroitin sulfate biosynthesis, CSS2B inhibits it. Analysis of transcription levels of CSS2 variants
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
35 ng/ml transforming growth factor-beta1 and epidermal growth factor upregulate chondroitin sulfate synthase-1 expression in microglias after brain injury
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knockdown of c-Jun and Sp1 decreases CHSY1 promoter activity, CHSY1 expression and sulfated glycosaminoglycan accumulation induced by TGF-beta
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TGF-beta induces up-regulation of CHSY1 in nucleus pulposus cells through MAPK signaling. TGF-beta increases CHSY1 expression and promoter activity in the nucleus pulposus partially through activation of canonical Smad signaling pathway
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D241K
-
inactive using UDP-N-acetyl-beta-D-galactosamine as substrate
D521K
-
inactive using UDP-N-glucuronic acid as substrate
D241K
-
inactive using UDP-N-acetyl-beta-D-galactosamine as substrate
-
D521K
-
inactive using UDP-N-glucuronic acid as substrate
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F362S
-
naturally occuring missense mutation of the CHSY1 activity-related, highly conserved residue, the mutant protein has a 56% decrease in GalNAcT-II activity and a 55% decrease in GlcAT-II activity compared to the wild-type
S126L
-
naturally occuring mutation, inactive mutant, the amount of chondroitin sulfate proteoglycans is reduced. In men, multiple sclerosis patients with S126L have a slower disease progression. This cSNP might be associated with the gender differences in clinical course of multiple sclerosis
D367A
-
site-directed mutagenesis, a soluble mutant is generated by replacing the first 36 amino acids of ChGn-2 D367A with a cleavable insulin signal sequence and the protein A IgG-binding domain, inactive mutant
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
diagnostics
-
chondroitin sulfate synthase 1 expression is associated with malignant potential of soft tissue sarcomas with myxoid substance. High expression of CHSY1 is significantly associated with poorer patient outcomes
medicine
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