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Information on EC 2.3.1.157 - glucosamine-1-phosphate N-acetyltransferase and Organism(s) Sulfurisphaera tokodaii and UniProt Accession Q975F9

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IUBMB Comments
The enzyme from several bacteria (e.g., Escherichia coli, Bacillus subtilis and Haemophilus influenzae) has been shown to be bifunctional and also to possess the activity of EC 2.7.7.23, UDP-N-acetylglucosamine diphosphorylase.
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This record set is specific for:
Sulfurisphaera tokodaii
UNIPROT: Q975F9
Word Map
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota, Archaea
The taxonomic range for the selected organisms is: Sulfurisphaera tokodaii
Synonyms
amino-sugar-1-P AcTase, amino-sugar-1-phosphate acetyltransferase, bifunctional GlmU protein, bifunctional protein GlmU, bifunctional UDP-N-acetylglucosamine pyrophosphorylase/glucosamine-1-phosphate N-acetyltransferase, galactosamine-1-phosphate acetyltransferase, GalN-1-P AcTase, GlcN-1-P acetyltransferase, GlcN-1-P AcTase, GlcNAc-1-P uridyltransferase, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
amino-sugar-1-P AcTase
299945
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amino-sugar-1-phosphate acetyltransferase
299945
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galactosamine-1-phosphate acetyltransferase
299945
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GalN-1-P AcTase
299945
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GlcN-1-P AcTase
299945
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glucosamine-1-phosphate acetyltransferase
299945
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ST0452
ST0452 protein
299945
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STK_04520
299945
locus name
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
acetyl-CoA + alpha-D-glucosamine 1-phosphate = CoA + N-acetyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
the multifunctional enzyme from Sulfolobus tokodaii {7} is also active with acetyl-CoA + alpha-D-galactosamine 1-phosphate (galactosamine-1-phosphate N-acetyltransferase), UTP + N-acetylglucosamine 1-phosphate (EC 2.7.7.23, UDP-N-acetylglucosamine diphosphorylase) and UTP + N-acetyl-alpha-D-galactosamine 1-phosphate (EC 2.7.7.83, UDP-N-acetylgalactosamine diphosphorylase)
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Acyl group transfer
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SYSTEMATIC NAME
IUBMB Comments
acetyl-CoA:alpha-D-glucosamine-1-phosphate N-acetyltransferase
The enzyme from several bacteria (e.g., Escherichia coli, Bacillus subtilis and Haemophilus influenzae) has been shown to be bifunctional and also to possess the activity of EC 2.7.7.23, UDP-N-acetylglucosamine diphosphorylase.
CAS REGISTRY NUMBER
COMMENTARY hide
9023-06-7
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9031-91-8
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SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
acetyl-CoA + alpha-D-galactosamine 1-phosphate
CoA + N-acetyl-alpha-D-galactosamine 1-phosphate
show the reaction diagram
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-
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?
acetyl-CoA + alpha-D-glucosamine 1-phosphate
CoA + N-acetyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
additional information
?
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the enzyme has multiple sugar-1-phosphate nucleotidylyltransferase, EC 2.7.7.37, and amino-sugar-1-phosphate acetyltransferase, EC 2.3.1.157, activities, overview. In addition to glucosamine-1-phosphate acetyltransferase activity, it possesses unique galactosamine-1-phosphate acetyltransferase activity. Also, the enzyme possesses GlcNAc-1-phosphate nucleotidylyltransferase, EC 2.7.7.23, and N-acetyl-D-galactosamine-1-phosphate uridyltransferase, EC 2.7.7.83, activities, as well as the expected glucose-1-phosphate thymidylyltransferase, EC 2.7.7.24, activity. The ST0452 protein can catalyze the acetylation of both GlcN-1-P and GalN-1-P, while GalN-1-P AcTase activity is not detected in bacterial enzymes
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NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
acetyl-CoA + alpha-D-galactosamine 1-phosphate
CoA + N-acetyl-alpha-D-galactosamine 1-phosphate
show the reaction diagram
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-
-
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?
acetyl-CoA + alpha-D-glucosamine 1-phosphate
CoA + N-acetyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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no metal ion required
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Ca2+
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2 mM, inhibits D-glucosamine-1-phosphate N-acetyltransferase activity
Co2+
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2 mM, inhibits D-glucosamine-1-phosphate N-acetyltransferase activity
Mg2+
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2 mM, inhibits D-glucosamine-1-phosphate N-acetyltransferase activity
Mn2+
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2 mM, inhibits D-glucosamine-1-phosphate N-acetyltransferase activity
Zn2+
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2 mM, inhibits D-glucosamine-1-phosphate N-acetyltransferase activity
additional information
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residues Tyr311, Lys337 and Lys340 plus C-terminal 11-residue region of the ST0452 protein enhance its GalN-1-P AcTase activity and suppress its GlcN-1-P AcTase activity, this function might be lost in bacterial enzymes
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.52 - 1.55
acetyl-CoA
0.66 - 1.71
alpha-D-galactosamine 1-phosphate
0.56 - 1.69
alpha-D-glucosamine 1-phosphate
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
12.6 - 69.7
alpha-D-galactosamine 1-phosphate
123.2 - 2311
alpha-D-glucosamine 1-phosphate
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
19.6 - 40.8
alpha-D-galactosamine 1-phosphate
211 - 1489
alpha-D-glucosamine 1-phosphate
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.35
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pH 7.5, 80°C, mutant enzyme H308A
3 - 8
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alpha-D-galactosamine 1-phosphate substrate, pH 7.5, 80°C, recombinant mutant D005 protein
23.1
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pH 7.5, 80°C, mutant enzyme N331A
29.6
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alpha-D-galactosamine 1-phosphate aubstrate, pH 7.5, 80°C, recombinant mutant D011 protein
47.6
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alpha-D-galactosamine 1-phosphate aubstrate, pH 7.5, 80°C, recombinant wild-type ST0452 protein
50
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pH 7.5, 80°C, wild-type enzyme enzyme
59.2
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pH 7.5, 80°C, mutant enzyme Y311A
67.4
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alpha-D-glucosamine 1-phosphate substrate, pH 7.5, 80°C, recombinant wild-type ST0452 protein
68.9
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pH 7.5, 80°C, mutant enzyme K337A
73.5
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pH 7.5, 80°C, mutant enzyme K340A
75.34
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pH 7.5, 80°C, substrates: acetyl-CoA + alpha-D-glucosamine 1-phosphate
323.5
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alpha-D-glucosamine 1-phosphate substrate, pH 7.5, 80°C, recombinant mutant D005 protein
1131
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alpha-D-glucosamine 1-phosphate aubstrate, pH 7.5, 80°C, recombinant mutant D011 protein
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
physiological function
additional information
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the C-terminal tail region of the ST0452 protein might be important for recognition of the multiple substrates for amino-sugar-1-P AcTase activity. The ST0452 protein contains only two Cys residues, it is unlikely that Cys-Cys bonds contribute to its thermostability. Residue Asn331 in the ST0452 protein is essential for the GalN-1-P AcTase activity, but it is much less important and not essential for the GlcN-1-P AcTase activity. The C-terminal residues of the ST0452 protein enhance the turnover rate of its GalN-1-P AcTase catalytic activity and slightly suppress substrate binding. Residue H308 is essential for both amino-sugar-1-P AcTase activities of the ST0452 protein
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
trimer
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the C-terminal domain of the ST0452 protein, with its LbetaH structure, appears to be essential for the formation of its trimeric form and, in turn, the high stability of the entire ST0452 protein
additional information
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the enzyme contains the the N-terminal nucleotidylyltransferase domain (residues 1–210) and the C-terminal acetyltransferase domain (residues 211–401), respectively. Comparisons of the crystal structures of the ST0452 protein, PDB ID GGO, and Escherichia coli protein EcGlmU2, PDB ID 2OI5, comparison with ST0452 mutant enzymes, overview. Despite the structural similarities between the N- and C-termini of the ST0452 protein and those of Escherichia coli EcGlmU, the thermostabilities of the two proteins differ greatly, as EcGlmU is a mesophilic enzyme. The structures of these proteins do not correlate directly with their thermostability. The distance between the GlcN-1-P AcTase and GlcNAc-1-P UTase catalytic centers is smaller in the ST0452 protein than the mesophilic bacterial GlmU
CRYSTALLIZATION/commentary
ORGANISM
UNIPROT
LITERATURE
sitting drop vapor diffusion method at 22°C, crystallization of the Y97N protein
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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
H308A
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site-directed mutagenesis, the mutation diminishes both amino-sugar-1-P AcTase activities of the ST0452 protein. The mutant shows 7.7% and 0.7% of wild-type GalN-1-P AcTase and GlcN-1-P AcTase activity, respetively; specific activity is 0.7% compared to the wild-type enzyme
K337A
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site-directed mutagenesis, the mutant enzyme shows slightly decreasing GalN-1-P AcTase activity and slightly increasing GlcN-1-P AcTase activity compared to the wild-type enzyme. The mutant shows 82.6% and 137.7% of wild-type GalN-1-P AcTase and GlcN-1-P AcTase activity, respetively
K340A
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site-directed mutagenesis, the mutant enzyme shows moderately decreasing GalN-1-P AcTase activity and moderately increasing GlcN-1-P AcTase activity compared to the wild-type enzyme. The mutant shows 63.3% and 147.1% of wild-type GalN-1-P AcTase and GlcN-1-P AcTase activity, respetively; specific activity is 147.1% compared to the wild-type enzyme
K377A
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specific activity is 137.7% compared to the wild-type enzyme
N331A
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site-directed mutagenesis, the mutant enzyme shows highly decreasing GalN-1-P AcTase activity and decreasing GlcN-1-P AcTase activity compared to the wild-type enzyme. The mutant shows 3.1% and 46.1% of wild-type GalN-1-P AcTase and GlcN-1-P AcTase activity, respetively; specific activity is 46.1% compared to the wild-type enzyme
T80S/Y97N
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the mutant enzyme shows 6.5times-higher activity, compared to that of the wild-type ST0452 protein, revealing that these two substituted residues function cooperatively to increase N-acetylglucosamine-1-phosphate uridyltransferase activity
Y311A
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site-directed mutagenesis, the mutant enzyme shows highly decreasing GalN-1-P AcTase activity and increasing GlcN-1-P AcTase activity compared to the wild-type enzyme. The mutant shows 3.3% and 118.4% of wild-type GalN-1-P AcTase and GlcN-1-P AcTase activity, respetively; specific activity is 118.4% compared to the wild-type enzyme
Y97N
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the mutant enzyme exhibits over 4 times higher N-acetylglucosamine-1-phosphate uridyltransferase activity, compared with that of the wild-type ST0452 protein. The three-dimensional structure of the Y97N protein is not changed by this substitution but the interactions with the substrate are slightly modified, which might cause the activity to increase. The crystal structure of the Y97N protein shows that positions 146 (Glu) and 80 (Thr) form interactions with GlcNAc, and an engineering strategy is applied to these residues to increase activity
additional information
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construction of expression vectors encoding a series of ST0452 C-terminal deletion mutants with hexahistidine tags at their C-termini, designated pST0452(DC005)H, pST0452(DC011)H, pST0452(DC021)H, pST0452(DC031)H, pST0452(DC041) H, pST0452(DC051)H, pST0452(DC071)H, pST0452 (DC121)H and pST0452(DC171)H. The deletion mutants retain the same tertiary structures as the wild-type ST0452 protein, but some show an altered thermostability, overview; glucosamine-1-phosphate acetyltransferase activity of C-terminal deletion mutants DC005 and DC011 (deletion of the C-terminal 5 or 11 residues of the ST0452 protein) are respectively, 4.8 and 16.8 times higher than that of the wild-type ST0452 protein. The mutant enzyme DC011 (deletion of the C-terminal 11 residues of the ST0452 protein) shows little thermal stability at 80°C. The C-terminal domain of the ST0452 protein, with its LbetaH structure, appears to be essential for the formation of its trimeric form and, in turn, the high stability of the entire ST0452 protein. The deletion mutant enzymes DC021, DC031, DC041, DC071 and DC121, are produced in an insoluble form or aggregated immediately after purification. Mutant enzymes DC051 and DC171 can be expressed in a soluble form. Mutant enzyme DC051 becomes completely insoluble after 5 min treatment at 60°C, while mutant enzyme DC171 is insoluble after 5 min treatment at 70 °C
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
80
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mutant enzyme DC005 shows the same thermostability as wild-type ST0452 protein, whereas mutant enzyme DC011 denatures and becomes insoluble by 5-min treatment at 80 °C. The C-terminal domain of the ST0452 protein, with its LbetaH structure, appears to be essential for the formation of its trimeric form and, in turn, the high stability of the entire ST0452 protein
PURIFICATION/commentary
ORGANISM
UNIPROT
LITERATURE
CLONED/commentary
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli strain BL21-Codon Plus(DE3)-RIL; gene ST0452, recombinant expression of C-terminally His6-tagged wild-type and mutant enzymes in Escherichia coli strain BL21-Codon Plus(DE3)-RIL
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expression of mutant enzymes in Escherichia coli
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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the increase in activity induced by some substitutions and truncations may be a useful feature that can be exploited for commercial application of this enzyme
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Zhang, Z.; Akutsu, J.; Kawarabayasi, Y.
Identification of novel acetyltransferase activity on the thermostable protein ST0452 from Sulfolobus tokodaii strain 7
J. Bacteriol.
192
3287-3293
2010
Sulfurisphaera tokodaii (Q975F9), Sulfurisphaera tokodaii, Sulfurisphaera tokodaii 7 (Q975F9)
Manually annotated by BRENDA team
Zhang, Z.; Shimizu, Y.; Kawarabayasi, Y.
Characterization of the amino acid residues mediating the unique amino-sugar-1-phosphate acetyltransferase activity of the archaeal ST0452 protein
Extremophiles
19
417-427
2015
Sulfurisphaera tokodaii (Q975F9), Sulfurisphaera tokodaii, Sulfurisphaera tokodaii 7 (Q975F9), Sulfurisphaera tokodaii DSM 16993 / JCM 10545 / NBRC 100140 / 7 (Q975F9)
Manually annotated by BRENDA team
Honda, Y.; Nakano, S.; Ito, S.; Dadashipour, M.; Zhang, Z.; Kawarabayasi, Y.
Improvement of ST0452 N-acetylglucosamine-1-phosphate uridyltransferase activity by the cooperative effect of two single mutations identified through structure-based protein engineering
Appl. Environ. Microbiol.
84
e002213-18
2018
Sulfurisphaera tokodaii (Q975F9), Sulfurisphaera tokodaii, Sulfurisphaera tokodaii DSM 16993 (Q975F9)
Manually annotated by BRENDA team
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