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Information on EC 3.5.1.26 - N4-(beta-N-acetylglucosaminyl)-L-asparaginase and Organism(s) Homo sapiens and UniProt Accession P20933
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3.5.1.26 N4-(beta-N-acetylglucosaminyl)-L-asparaginaseIUBMB Comments
Acts only on asparagine-oligosaccharides containing one amino acid, i.e., the asparagine has free alpha-amino and alpha-carboxyl groups [cf. EC 3.5.1.52, peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase]
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Word Map
- 3.5.1.26
- lysosomal
- aspartylglucosaminuria
- lepidoptera
-
mitogenome
-
protein-coding
-
microsatellite-like
-
attta
- poly-a
- butterfly
-
a+t-rich
- glycoasparagines
- nymphalidae
- aspartylglucosamine
- dihydrouridine
-
trnaseragn
-
autoproteolysis
- meningosepticum
- hesperiidae
- papilionidae
-
lrrna
- pieridae
-
non-finnish
- glcnac-asn
-
clover-leaf
- medicine
- lycaenidae
- trnamet
- diagnostics
The taxonomic range for the selected organisms is: Homo sapiens
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota, Archaea
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota, Archaea
Synonyms
glycosylasparaginase, aspartylglucosaminidase, ataga, glycoasparaginase, amidase-2, 1-aspartamido-beta-n-acetylglucosamine amidohydrolase, n-aspartyl-beta-glucosaminidase, aspartylglucosylaminase, amidase-3, n4-(n-acetyl-beta-glucosaminyl)-l-asparagine amidase, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
AGA
aspartylglucosaminidase
aspartylglucosylaminase
-
-
-
-
aspartylglucosylamine deaspartylase
-
-
-
-
aspartylglycosylamine amidohydrolase
-
-
-
-
beta-aspartylglucosylamine amidohydrolase
-
-
-
-
glucosylamidase
-
-
-
-
N-aspartyl-beta-glucosaminidase
-
-
-
-
N4-(N-acetyl-beta-glucosaminyl)-L-asparagine amidase
-
-
-
-
additional information
belongs to the N-terminal nucleophile hydrolase superfamily
AGA
-
-
-
-
aspartylglucosaminidase
-
-
-
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O = N-acetyl-beta-D-glucosaminylamine + L-aspartate
mechanism
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O = N-acetyl-beta-D-glucosaminylamine + L-aspartate
mechanism
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
carboxylic acid amide hydrolysis
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine amidohydrolase
Acts only on asparagine-oligosaccharides containing one amino acid, i.e., the asparagine has free alpha-amino and alpha-carboxyl groups [cf. EC 3.5.1.52, peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase]
CAS REGISTRY NUMBER
COMMENTARY
9075-24-5
-
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
aspartic acid beta-(4-nitroanilide) + H2O
4-nitroaniline + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
N-acetyl-beta-D-glucosaminylamine + L-aspartate
Thr-206 is the N-terminal nucleophile that acts as catalytic residue, Thr-206 is stabilized by hydrogen bonds from Ser-72 and Thr-224, enzyme structure
-
-
?
N4-(beta-N-acetylglucosaminyl)-L-asparagine + H2O
N-acetyl-D-glucosaminylamine + L-aspartate
-
-
-
?
2-acetamido-1-beta-(L-aspartamido)-1,2-dideoxy-beta-D-glucose + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
L-asparagine + H2O
L-aspartate + NH3
-
21% activity compared to N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine as substrate
-
r
L-aspartic acid beta-(7-amido-4-methylcoumarin) + H2O
7-amino-4-methylcoumarin + L-aspartate
L-aspartic acid-b-7-amido-4-methylcoumarin + H2O
7-amino-4-methylcoumarin + L-aspartate
-
-
-
-
?
N4-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-L-asparagine + H2O
2-acetamido-2-deoxy-beta-D-glucopyranosylamine + L-asparagine
-
catalyzes the hydrolysis of the N-glycosylic bond
-
-
?
N4-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-L-asparagine + H2O
?
-
catabolism of N-linked oligosaccharides
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
N-acetyl-beta-D-glucosaminylamine + L-aspartate
-
enzyme catalyzes the hydrolysis of the N-glycosylic bond between asparagine and N-acetylglucosamine, mechanism involving formation of a tetra-hedral high-energy intermediate, presence of a second binding site that may recognize substituted acetamido groups
-
-
?
2-acetamido-1-beta-(L-aspartamido)-1,2-dideoxy-beta-D-glucose + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
2-acetamido-1-beta-(L-aspartamido)-1,2-dideoxy-beta-D-glucose + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
2-acetamido-1-beta-(L-aspartamido)-1,2-dideoxy-beta-D-glucose + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
complex enzyme system in tissue homogenates
-
?
L-aspartic acid beta-(7-amido-4-methylcoumarin) + H2O
7-amino-4-methylcoumarin + L-aspartate
-
-
-
?
L-aspartic acid beta-(7-amido-4-methylcoumarin) + H2O
7-amino-4-methylcoumarin + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
involved in degradation of glycoproteins
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
involved in degradation of glycoproteins
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
involved in degradation of glycoproteins
-
r
additional information
?
-
aspartylglucosaminuria, a lysosomal storage disease caused by mutation L15R, is enriched in the Finnish population
-
-
?
additional information
?
-
-
aspartylglucosaminuria, a lysosomal storage disease caused by mutation L15R, is enriched in the Finnish population
-
-
?
additional information
?
-
processing and activation of aspartylglucosaminidase by autocatalytic cleavage, overview
-
-
?
additional information
?
-
-
processing and activation of aspartylglucosaminidase by autocatalytic cleavage, overview
-
-
?
additional information
?
-
-
enzyme catalyzes the hydrolysis of the N-glycosylic bond between asparagine and N-acetylglucosamine in the catabolism of N-linked glycoproteins
-
-
?
additional information
?
-
-
enzyme catalyzes the hydrolysis of a variety of asparagine and aspartyl compounds containing a free alpha-carboxyl group and a free alpha-amino group, study of enzyme specificity with 14 substrate analogues where the structure of the aspartyl group is changed, the alpha-carboxyl group is necessary for enzyme activity, but not the alpha-amino group, whose position acts as an anchor in the binding site for the substrate, no substrate: N-acetyl-D-glucosamine, intramolecular autoproteolytic activation reaction
-
-
?
additional information
?
-
-
enzyme level in plasma is elevated in congenital disorders of glycosylation type I, CDG I, a defect in biosynthesis or processing of O-linked glycans, overview
-
-
?
additional information
?
-
-
glycosylasparaginase hydrolyzes the N-glycosidic carbohydrate-to-protein linkage region, aspartylglucosamine, to L-aspartic acid and L-amino-N-acetylglucosamine through a reaction mechanism similar to L-asparaginase
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
N4-(beta-N-acetylglucosaminyl)-L-asparagine + H2O
N-acetyl-D-glucosaminylamine + L-aspartate
-
-
-
?
L-asparagine + H2O
L-aspartate + NH3
-
21% activity compared to N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine as substrate
-
r
N4-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-L-asparagine + H2O
?
-
catabolism of N-linked oligosaccharides
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
-
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
involved in degradation of glycoproteins
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
involved in degradation of glycoproteins
-
?
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O
2-acetamido-1-amino-1,2-dideoxy-beta-D-glucose + L-aspartate
-
involved in degradation of glycoproteins
-
r
additional information
?
-
aspartylglucosaminuria, a lysosomal storage disease caused by mutation L15R, is enriched in the Finnish population
-
-
?
additional information
?
-
-
aspartylglucosaminuria, a lysosomal storage disease caused by mutation L15R, is enriched in the Finnish population
-
-
?
additional information
?
-
-
enzyme catalyzes the hydrolysis of the N-glycosylic bond between asparagine and N-acetylglucosamine in the catabolism of N-linked glycoproteins
-
-
?
additional information
?
-
-
enzyme level in plasma is elevated in congenital disorders of glycosylation type I, CDG I, a defect in biosynthesis or processing of O-linked glycans, overview
-
-
?
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
3-phosphono-DL-2-aminopropionic acid
-
phosphono transition state mimic, competitive inhibitor
N-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)bromoacetamide
-
substrate analogue, noncompetitive inhibitor
N-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)chloroacetamide
-
substrate analogue, noncompetitive inhibitor
N1-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)glycinamide
-
substrate analogue, noncompetitive inhibitor
SDS
-
decreasing activity only after heating above 60°C in the presence of 0.1-0.5% SDS, w/v
5-Diazo-4-oxo-L-norvaline
-
irreversible inhibition, Ki: 0.08 mM, competitively protected by its natural substrate
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
intramolecular autoproteolytic activation with Gly-258 playing an important structural role, molecular activation mechanism
-
additional information
-
intramolecular autoproteolytic activation with Gly-258 playing an important structural role, molecular activation mechanism
-
additional information
processing and activation of aspartylglucosaminidase by autocatalytic cleavage, overview
-
additional information
-
processing and activation of aspartylglucosaminidase by autocatalytic cleavage, overview
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.143
2-acetamido-1-beta-(L-aspartamido)-1,2-dideoxy-D-glucose
pH 7, 37°C, purified wild-type AGA
0.444
2-acetamido-1-beta-(L-aspartamido)-1,2-dideoxy-D-glucose
pH 7, 37°C, wild-type AGA
0.09
N4-(beta-N-acetylglucosaminyl)-L-asparagine
recombinant wild-type enzyme, pH 7.5, 37°C
0.166
N4-(beta-N-acetylglucosaminyl)-L-asparagine
mutant T203I, pH 7.5, 37°C
0.26
N4-(beta-N-acetylglucosaminyl)-L-asparagine
mutant T99K, pH 7.5, 37°C
0.16
N4-(beta-N-acetyl-D-glucosaminyl)-L-asparagine
-
at 37°C and pH 7.7, pH- and temperature dependend
additional information
additional information
kinetics of mutant T203I and wild-type enzymes, the specificity constant (kcat/Km) of mutant T203I is decreased by more than 300fold when compared to the wild-type enzyme
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.08
N4-(beta-N-acetylglucosaminyl)-L-asparagine
mutant T203I, pH 7.5, 37°C
5.93
N4-(beta-N-acetylglucosaminyl)-L-asparagine
mutant T99K, pH 7.5, 37°C
14.18
N4-(beta-N-acetylglucosaminyl)-L-asparagine
recombinant wild-type enzyme, pH 7.5, 37°C
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.482
N4-(beta-N-acetylglucosaminyl)-L-asparagine
mutant T203I, pH 7.5, 37°C
22.8
N4-(beta-N-acetylglucosaminyl)-L-asparagine
mutant T99K, pH 7.5, 37°C
157.6
N4-(beta-N-acetylglucosaminyl)-L-asparagine
recombinant wild-type enzyme, pH 7.5, 37°C
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.64
N-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)chloroacetamide
-
pH 5.8, 37°C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
0.72
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
AGA maturation, activation cleavage of the dimerized AGA precursors into the N-terminal alpha- and the C-terminal beta-subunits takes place in the endoplasmic reticulum
additional information
additional information
L15R causes aspartylglucosaminuria and affects translocation of aspartylglucosaminidase
-
additional information
-
L15R causes aspartylglucosaminuria and affects translocation of aspartylglucosaminidase
-
additional information
the pro-enzyme contains a 23 residues signal peptide
-
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
evolution
belongs to the group of so-called N-terminal nucleophile (NTN) hydrolases. The members of the NTN hydrolase family, which in addition to AGA also include, e.g., the proteasome beta-subunit and penicillin acylase, show very little similarity at the amino acid sequence level, but they exhibit a highly similar folded structure
malfunction
physiological function
additional information
malfunction
aspartylglucosaminuria (AGU) is an inherited disease caused by mutations in a lysosomal amidase called aspartylglucosaminidase (AGA) or glycosylasparaginase. This disorder results in an accumulation of glycoasparagines in the lysosomes of virtually all cell types, with severe clinical symptoms affecting the central nervous system, skeletal abnormalities, and connective tissue lesions. Many AGU mutations remain as dimers but cannot undergo autoproteolysis and thus lack amidase activity for digesting glycoasparagines
malfunction
defects in the AGA gene result in a lysosomal storage disorder, aspartylglucosaminuria (AGU), that manifests mainly as progressive mental retardation. A number of AGU missense mutations have been identified that result in reduced AGA activity. Human variants contain either Ser or Thr in position 149, and Thr149 AGA, which is the rare variant, can be considered as a neutral or benign variant. AGU mutations result in reduced AGA activity in patient cells. Mutations in the AGA gene result in aspartylglucosaminuria (AGU, OMIM 208400), a lysosomal storage disorder that is characterized by progressive loss of intellectual capabilities and some skeletal abnormalities. AGU patients are born seemingly normal, but the progressive course of the disease manifests in, e.g., developmental delay, loss of speech and coarse facial features early in childhood. In adulthood, most AGU patients are severely retarded and require special care
malfunction
deficiency of the enzyme causes accumulation of glycoasparagines in lysosomes of cells, resulting in a genetic condition called aspartylglucosaminuria (AGU). AGU is a lysosomal storage disorder caused by defects of the hydrolase glycosylasparaginase. AGU mutations impair autoproteolysis of GA precursor and/or impair its hydrolase activity in lysosomes. This deficiency results in progressive mental decline of the patients, and leads to a lifelong condition affecting patient's appearance, cognition, adaptive skills, physical growth, personality, anatomical structure, and health. Early indicators of AGU include growth spurts in infants and the development of macrocephalia associated to hernias and respiratory infections
physiological function
aspartylglucosaminidase (AGA) is a lysosomal hydrolase that participates in the breakdown of glycoproteins
physiological function
the enzyme is involved in protein degradation by cleaving Asn-linked glycoproteins in lysosomes. During the metabolic turnover of Asn-linked glycoproteins, autocleaved enzyme hydrolyzes glycoasparagine N4-(beta-N-acetylglucosaminyl)-L-asparagine (NAcGlc-Asn) that connects a carbohydrate to the side chain of an asparagine
physiological function
the enzyme is is involved in protein degradation by catabolizing Asn-linked glycoproteins in lysosomes
additional information
AGA belongs to the group of so-called N-terminal nucleophile (NTN) hydrolases, as the free alpha-amino group of Thr206 is involved in the catalysis as the base, whereas the OH group of Thr206 functions as a nucleophile during the catalysis
additional information
-
AGA belongs to the group of so-called N-terminal nucleophile (NTN) hydrolases, as the free alpha-amino group of Thr206 is involved in the catalysis as the base, whereas the OH group of Thr206 functions as a nucleophile during the catalysis
additional information
structure comparison of mutant T203I with the wild-type enzyme
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). ASPG_HUMAN
346
0
37208
Swiss-Prot
Secretory Pathway (Reliability: 1)
PDB
SCOP
CATH
UNIPROT
ORGANISM
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
18000
24000
25000
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
heterotetramer
dimer
tetramer
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
proteolytic modification
glycoprotein
proteolytic modification
-
intramolecular autoproteolytic activation reaction
proteolytic modification
intramolecular autoproteolytic activation with Gly-258 playing an important structural role, molecular activation mechanism, dimerization and correct folding of the AGA precursor, activation cleavage of the dimerized AGA precursors into the N-terminal alpha- and the C-terminal beta-subunits takes place in the endoplasmic reticulum and results in the tetrameric, enzymically active (alpha,beta)2 molecule
proteolytic modification
the enzyme contains a subcellular targeting signal sequence
proteolytic modification
processing and activation of aspartylglucosaminidase by autocatalytic cleavage, overview. AGA is synthesized in the endoplasmic reticulum as a 346 amino acid (aa) polypeptide from which 23 residues of the signal peptide are removed. Very soon after synthesis in the endoplasmic reticulum, two AGA precursors homodimerize, inducing an autocatalytic cleavage of both precursors N-terminally to Thr206 into 27 kDa pro-alpha and 17 kDa subunits. After transport to lysosomes, the pro-alpha is C-terminally cleaved into 24 kDa mature alpha-subunit, whereas processing of the beta-subunit gives rise to the 14 kDa beta'-subunit. Neither of these lysosomal processing steps displays an effect on the enzyme activity
proteolytic modification
the enzyme is processed through autocatalytic cleavage. A subsequent autoprocessing results in a main-chain cleavage at the P-loop by a self-catalyzed peptide bond rearrangement via an N -> O acyl shift. This autoproteolysis event results in an active form of the hydrolase with separate alpha- and beta-subunits
proteolytic modification
the enzyme is synthesized as a single-chain precursor that requires an intramolecular autoprocessing to form a mature amidase
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
purified recombinant enzyme mutant T203I free and in complex with N4-(beta-N-acetylglucosaminyl)-L-asparagine, hanging drop vapor diffusion technique, mixing of 2mg/ml protein solution with reservoir oslution containing 0.2 M NaCl, 0.1 M bis-Tris, pH 6.5, and 25% PEG 3350, cryoprotection in solution containing 100 mM Tris-HCl, pH 8.0, and 20% glycerol, X-ray diffraction structure determination and analysis, molecular replacement using the structure of the wild-type enzyme (PDB ID 2GAW) as the starting model
purified recombinant enzyme mutant T99K free and in complex with substrate N4-(beta-N-acetylglucosaminyl)-L-asparagine, hanging drop vapor diffusion technique, mixing of 2mg/ml protein solution with reservoir oslution containing 0.2 M NaCl, 0.1 M bis-Tris, pH 6.5, and 25% PEG 3350, cryoprotection in solution containing 100 mM Tris-HCl, pH 8.0, and 20% glycerol, X-ray diffraction structure determination and analysis at 1.5 A resolution, molecular replacement using the structure of the wild-type enzyme (PDB ID 2GAW) as the starting model
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
C140S
the substitution is the causative mutation for enzyme deficiency. In addition to preventing the disulfide bond formation between C140 and C156, the C140S substitution also causes destabilization of the unique/longer loop structure in the human sequence and thus prevent dimerization of GA essential for autoproteolytic activation
D200A
87% of wild-type activity, study of folding, transport and catalytic kinetics of mutant AGA
D201A
93% of wild-type activity, study of folding, transport and catalytic kinetics of mutant AGA
D70A
44% of wild-type activity, study of folding, transport and catalytic kinetics of mutant AGA
G172D
G203D
G226A
inactive mutant, study of folding, transport and catalytic kinetics of mutant AGA
G226D
G258A
inactive mutant, study of folding, transport and catalytic kinetics of mutant AGA
K230A
86% of wild-type activity, study of folding, transport and catalytic kinetics of mutant AGA
L15R
naturally occuring L15R, mutating the signal sequence, causes aspartylglucosaminuria and affects translocation of aspartylglucosaminidase
N225A
45% of wild-type activity, study of folding, transport and catalytic kinetics of mutant AGA
R138Q
the single mutation does not affect either the enzyme's autoproteolysis or its hydrolase activity
R138Q/C140S
naturally occuring mutation in Finnish population causing aspartylglucosaminuria (AGU). Due to a founder effect, AGU incidence in Finland is unexcelled, with one major allele (denoted AGUFIN) found in 98% of the AGU patients. The AGUFIN allele carries the two concurrent substitutions R138Q andC140S
R161Q/C163S
naturally occuring mutation, the AGUFin-major mutation is a combination of two missense mutations, abolishing a disulfide bond and destabilizing the AGA structure. The pathogenic C163S substitution is always combined with a functionally neutral Arg161Gln substitution. Mutations in the AGA gene result in aspartylglucosaminuria (AGU, OMIM 208400), a lysosomal storage disorder that is characterized by progressive loss of intellectual capabilities and some skeletal abnormalities. AGU patients are born seemingly normal, but the progressive course of the disease manifests in, e.g. developmental delay, loss of speech and coarse facial features early in childhood. In adulthood, most AGU patients are severely retarded and require special care
S238A
40% of wild-type activity, study of folding, transport and catalytic kinetics of mutant AGA
T122K
T203I
T234I
T257I
naturally occuring mutation in Canadian population causing aspartylglucosaminuria (AGU), the mutant lacks the signal peptide
T33A
48% of wild-type activity, study of folding, transport and catalytic kinetics of mutant AGA
T99K
H124R
-
reduced dimerization of the precursor polypeptide, total secretion into the medium
H124W
-
reduced dimerization of the precursor polypeptide, total secretion into the medium
N38D
-
30% of wild-type activity, proper processing to its mature lysosomal form
T257A
T310A
-
less than 10% of wild-type activity, more properly cleaved form than N308D
additional information
G172D
naturally occuring mutation in Finnish population causing aspartylglucosaminuria (AGU)
G172D
site-directed mutagenesis, the mutation causes a local conformational change, which in turn disrupts the requisite autoprocessing step to generate metabolically functional mature hydrolase
G203D
naturally occuring mutation in Canadian population, causing aspartylglucosaminuria (AGU)
G203D
naturally occuring mutation in US-American population causing aspartylglucosaminuria (AGU)
G226D
naturally occuring mutation in Canadian population causing aspartylglucosaminuria (AGU)
G226D
naturally occuring mutation in Canadian population, causing aspartylglucosaminuria (AGU)
T122K
naturally occuring mutation in Canadian population causing aspartylglucosaminuria (AGU), the mutant lacks the signal peptide
T122K
naturally occuring mutation in US-American population causing aspartylglucosaminuria (AGU)
T203I
naturally occuring mutation in Finnish population causing aspartylglucosaminuria (AGU), the mutant lacks the signal peptide
T203I
site-directed mutagenesis, corresponds to mutation T234I, the replacement of the conserved threonine with an isoleucine has negative impacts on both KM and, to a greater extent, kcat of hydrolase activity, the mutant is almost inactive. Structure comparison of mutant T203I with the wild-type enzyme. Mutant T203I is capable of autoprotolysis. In the T203I-substrate complex model, all the previously identified substrate binding residues (W11, F13, S50, T152, R180, D183, G204, G206) are in close contact distances from the substrate model, except the mutated residue Ile203
T234I
naturally occuring mutation in Canadian population,located at the rim of the substrate binding site , causing aspartylglucosaminuria (AGU). The mutatnt enzyme shows reduced autoprocessing capability compared to wild-type
T234I
naturally occuring mutation in US-American population causing aspartylglucosaminuria (AGU), the mutant lacks the signal peptide
T99K
naturally occuring mutation in Finnish population causing aspartylglucosaminuria (AGU), the mutant lacks the signal peptide
T99K
naturally occuring mutation in US-American population causing aspartylglucosaminuria (AGU). This T99K model enzyme still has autoprocessing capacity to generate a mature form, its amidase activity to digest glycoasparagines remains low. A molecular clamp capable of fixing local disorders at the dimer interface might be able to rescue the deficiency of this AGU variant. The mutant lacks the signal peptide, but shows relatively high amidase activity, about 75% compared to wild-type. T99K has its substratebinding site fully opened through autoproteolysis and is ready to accommodate the substrate NAcGlc-Asn
additional information
aspartylglucosaminuria-causing mutations, most of them lead to defective molecular maturation of AGA, effects of targeted amino acid substitutions on the activation process of AGA
additional information
-
aspartylglucosaminuria-causing mutations, most of them lead to defective molecular maturation of AGA, effects of targeted amino acid substitutions on the activation process of AGA
additional information
construction of a model enzyme corresponding to a Finnish AGU allele, the naturally occuring T234I mutant variant. The Finnish variant is capable of a slow autoprocessing to generate detectible hydrolyzation activity of the natural substrate of the enzyme. Determination of a 1.6 A-resolution structure of the Finnish AGU model and construction of an enzyme-substrate complex to provide a structural basis for analyzing the negative effects of the point mutation on Km and kcat of the mature enzyme, overview
additional information
single nucleotide polymorphism rs2228119 (NM_000027.3:c.446C>G - p.(Thr149Ser)) results in amino acid variation Ser vs. Thr at position 149 (base and amino acid variation in red) of the human AGA enzyme. Functional analysis of the Ser149/Thr149 variants of human aspartylglucosaminidase and optimization of the coding sequence for protein production. Codon-optimized versions of the two variants are expressed at significantly higher levels than AGA with the natural codon-usage. The second most common allele in Finland is a 2 bp deletion called AGUFin-minor (NM_000027.3; c.199_200del - p.Glu67fc*3). Genotype frequency of single nucleotide polymorphism (SNP) rs2228119 in various populations
additional information
-
single nucleotide polymorphism rs2228119 (NM_000027.3:c.446C>G - p.(Thr149Ser)) results in amino acid variation Ser vs. Thr at position 149 (base and amino acid variation in red) of the human AGA enzyme. Functional analysis of the Ser149/Thr149 variants of human aspartylglucosaminidase and optimization of the coding sequence for protein production. Codon-optimized versions of the two variants are expressed at significantly higher levels than AGA with the natural codon-usage. The second most common allele in Finland is a 2 bp deletion called AGUFin-minor (NM_000027.3; c.199_200del - p.Glu67fc*3). Genotype frequency of single nucleotide polymorphism (SNP) rs2228119 in various populations
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
60
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
to homogeneity, chromatography techniques
-
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
DNA sequence determination and analysis, overexpression of mutant L15R in BHK and COS-1 cells, low expression level in endoplasmic reticulum, the recombinant active enzyme does not reach the lysosomes
gene AGA, genotyping, recombinant expression of codon-optimized genes encoding enzyme variants S149 and T149 in HEK-293T and HeLa cells, the Thr149 variant exhibits a slightly higher expression level than the Ser149 variant. Recombinant expression of AGA variants in transgenic patient fibroblasts, both wild-type and AGUFin-major mutant type. The transfection with the AGA variants improves the lysosomal morphology in AGU fibroblasts, low transfection efficiency
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
medicine
diagnostics
medicine
medicine
deficient AGA activity results in a lysosomal storage disease, aspartylglucosaminuria, resulting in progressive neurodegeneration, most of disease-causing mutations lead to defective molecular maturation of AGA
medicine
use of codon-optimized AGA may be beneficial for the therapy options in treatment of the lysosomal storage disorder aspartylglucosaminuria (AGU)
diagnostics
-
elevation of plasma aspartylglucosaminidase is a useful marker for the congenital disorders of glycosylation type I, CDG I
medicine
-
fluorometric assay in blood samples allows specific detection of the enzyme defect in aspartylglycosaminuria patients
medicine
-
enzyme deficiency or its absence gives rise to aspartylglycosaminuria, an autosomal recessive inherited disorder that results in the accumulation of glycoasparagines in lysosomes, most common disorder of glycoprotein metabolism
medicine
-
enzyme deficiency or its absence gives rise to aspartylglycosaminuria, the most common disorder of glycoprotein metabolism
medicine
-
human glycosylasparaginase is a potential anticancer agent to induce apoptosis in L-asparagine-dependent cancer cells
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Noronkoski, T.; Stoineva, I.B.; Petkov, D.D.; Mononen, I.
Recombinant human glycosylasparaginase catalyzes hydrolysis of L-asparagine
FEBS Lett.
412
149-152
1997
Homo sapiens
Saarela, J.; Laine, M.; Tikkanen, R.; Oinonen, C.; Jalanko, A.; Rouvinen, J.; Peltonen, L.
Activation and oligomerization of aspartylglucosaminidase
J. Biol. Chem.
273
25320-25328
1998
Homo sapiens
Tikkanen, R.; Riikonen, A.; Oinonen, C.; Rouvinen, J.; Peltonen, L.
Functional analyses of active site residues of human lysosomal aspartylglucosaminidase: implications for catalytic mechanism and autocatalytic activation
EMBO J.
15
2954-2960
1996
Homo sapiens
Park, H.; Vettese-Dadey, M.; Aronson, N.N.
Glycosylation and phosphorylation of lysosomal glycosylasparaginase
Arch. Biochem. Biophys.
328
73-77
1996
Homo sapiens
Mononen, I.; Mononen, T.; Ylikangas, P.; Kaartinen, V.; Savolainen, K.
Enzymatic diagnosis of aspartylglycosaminuria by fluorometric assay of glycosylasparaginase in serum, plasma, or lymphocytes
Clin. Chem.
40
385-388
1994
Homo sapiens
Enomaa, N.; Heiskanen, T.; Halila, R.; Sormunen, R.; Seppl, R.; Vihinen, M.; Peltonen, L.
Human aspartylglucosaminidase
Biochem. J.
286
613-618
1992
Homo sapiens
-
Rip, J.W.; Coulter-Mackie, M.B.; Rupar, C.A.; Gordon, B.A.
Purification and structure of human liver aspartylglucosaminidase
Biochem. J.
288
1005-1010
1992
Homo sapiens
Tollersrud, O.K.; Aronson, N.
Comparison of liver glycosylasparaginases from six vertebrates
Biochem. J.
282
891-897
1992
Bos taurus, Gallus gallus, Homo sapiens, Mus musculus, Rattus sp., Sus scrofa
Kaartinen, V.
Glycoasparaginase in human urine
Biochim. Biophys. Acta
1097
28-30
1991
Homo sapiens
Kaartinen, V.; Williams, J.C.; Tomich, J.; Yates, J.R.; Hood, L.E.; Mononen, I.
Glycosasparaginase from human leukocytes
J. Biol. Chem.
266
5860-5869
1991
Homo sapiens
Halila, R.; Baumann, M.; Ikonen, E.; Enomaa, N.; Peltonen, L.
Human leucocyte aspartylglucosaminidase
Biochem. J.
276
251-256
1991
Homo sapiens
McGovern, M.M.; Aula, P.; Desnick, R.J.
Purification and properties of human hepatic aspartylglucosaminidase
J. Biol. Chem.
258
10743-10747
1983
Homo sapiens
Dugal, B.
Effect of different compounds on 1-aspartamido-beta-N-acetylglucosamine amidohydrolase from human liver
Biochem. J.
171
799-802
1978
Homo sapiens
Dugal, B.; Stromme, J.
Purification and some properties of 1-aspartamido-beta-N-acetylglucosamine amidohydrolase from human liver
Biochem. J.
165
497-502
1977
Homo sapiens
Dugal, B.
Measurement of 1-aspartamido-beta-N-acetylglucosamine amidohydrolase activity in human tissues
Biochem. J.
163
9-14
1977
Homo sapiens
Risley, J.M.; Huang, D.H.; Kaylor, J.J.; Malik, J.J.; Xia, Y.Q.; York, W.M.
Glycosylasparaginase activity requires the alpha-carboxyl group, but not the alpha-amino group, on N(4)-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-L-asparagine
Arch. Biochem. Biophys.
391
165-170
2001
Homo sapiens
Saarela, J.; Oinonen, C.; Jalanko, A.; Rouvinen, J.; Peltonen, L.
Autoproteolytic activation of human aspartylglucosaminidase
Biochem. J.
378
363-371
2004
Homo sapiens (P20933), Homo sapiens
Risley, J.M.; Huang, D.H.; Kaylor, J.J.; Malik, J.J.; Xia, Y.Q.
Glycosylasparaginase inhibition studies: competitive inhibitors, transition state mimics, noncompetitive inhibitors
J. Enzyme Inhib.
16
269-274
2001
Homo sapiens
Saarela, J.; von Schantz, C.; Peltonen, L.; Jalanko, A.
A novel aspartylglucosaminuria mutation affects translocation of aspartylglucosaminidase
Hum. Mutat.
24
350-351
2004
Homo sapiens (P20933), Homo sapiens
Jackson, M.; Clayton, P.; Grunewald, S.; Keir, G.; Mills, K.; Mills, P.; Winchester, B.; Worthington, V.; Young, E.
Elevation of plasma aspartylglucosaminidase is a useful marker for the congenital disorders of glycosylation type I (CDG I)
J. Inherit. Metab. Dis.
28
1197-1198
2005
Homo sapiens
Mills, K.; Mills, P.; Jackson, M.; Worthington, V.; Beesley, C.; Mann, A.; Clayton, P.; Grunewald, S.; Keir, G.; Young, L.; Langridge, J.; Mian, N.; Winchester, B.
Diagnosis of congenital disorders of glycosylation type-I using protein chip technology
Proteomics
6
2295-2304
2006
Homo sapiens
Kelo, E.; Noronkoski, T.; Mononen, I.
Depletion of L-asparagine supply and apoptosis of leukemia cells induced by human glycosylasparaginase
Leukemia
23
1167-1171
2009
Homo sapiens
Pande, S.; Bizilj, W.; Guo, H.C.
Biochemical and structural insights into an allelic variant causing the lysosomal storage disorder - aspartylglucosaminuria
FEBS Lett.
592
2550-2561
2018
Homo sapiens (P20933)
Banning, A.; Koenig, J.F.; Gray, S.J.; Tikkanen, R.
Functional analysis of the Ser149/Thr149 variants of human aspartylglucosaminidase and optimization of the coding sequence for protein production
Int. J. Mol. Sci.
18
706
2017
Aurelia aurita, Homo sapiens (P20933), Homo sapiens
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