Information on EC 2.7.1.162 - N-acetylhexosamine 1-kinase

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The enzyme appears in viruses and cellular organisms

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EC NUMBER
COMMENTARY hide
2.7.1.162
-
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RECOMMENDED NAME
GeneOntology No.
N-acetylhexosamine 1-kinase
-
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + N-acetyl-D-hexosamine = ADP + N-acetyl-alpha-D-hexosamine 1-phosphate
show the reaction diagram
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Phosphorylation
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SYSTEMATIC NAME
IUBMB Comments
ATP:N-acetyl-D-hexosamine 1-phosphotransferase
This enzyme is involved in the lacto-N-biose I/galacto-N-biose degradation pathway in the probiotic bacterium Bifidobacterium longum. Differs from EC 2.7.1.157, N-acetylgalactosamine kinase, as it can phosphorylate both N-acetylgalactosamine and N-acetylglucosamine at similar rates. Also has some activity with N-acetyl-D-mannosamine, D-talose and D-mannose as substrate. ATP can be replaced by GTP or ITP but with decreased enzyme activity. Requires a divalent cation, with Mg2+ resulting in by far the greatest stimulation of enzyme activity.
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CAS REGISTRY NUMBER
COMMENTARY hide
959459-09-7
protein sequence
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GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
metabolism
additional information
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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
ATP + 2-(acetylamino)-2-deoxy-D-allopyranose
ADP + 2-(acetylamino)-2-deoxy-D-allopyranose 1-phosphate
show the reaction diagram
22% phosphorylation yield
-
-
?
ATP + 2-deoxy-2-azido-D-mannose
ADP + 2-deoxy-2-azido-alpha-D-mannose 1-phosphate
show the reaction diagram
ATP + 2-deoxy-2-fluoro-D-mannose
ADP + 2-deoxy-2-fluoro-alpha-D-mannose 1-phosphate
show the reaction diagram
ATP + 2-deoxy-D-glucose
ADP + 2-deoxy-alpha-D-glucose 1-phosphate
show the reaction diagram
ATP + 4- deoxy-N-acetyl-D-glucosamine
ADP + 4-deoxy-N-acetyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
-
50% phosphorylation yield
-
-
?
ATP + 4-deoxy-4-azido-N-acetyl-D-galactosamine
ADP + 4-deoxy-4-azido-N-acetyl-alpha-D-galactosamine 1-phosphate
show the reaction diagram
-
73% phosphorylation yield
-
-
?
ATP + 4-deoxy-D-mannose
ADP + 4-deoxy-alpha-D-mannose 1-phosphate
show the reaction diagram
ATP + 6-deoxy-6-azido-N-acetyl-D-galactosamine
ADP + 6-deoxy-6-azido-N-acetyl-alpha-D-galactosamine 1-phosphate
show the reaction diagram
-
42% phosphorylation yield
-
-
?
ATP + 6-deoxy-6-azido-N-acetyl-D-glucosamine
ADP + 6-deoxy-6-azido-N-acetyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
ATP + 6-deoxy-6-methyl-N-acetyl-D-glucosamine
ADP + 6-deoxy-6-methyl-N-acetyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
ATP + 6-deoxy-N-acetyl-D-galactosamine
ADP + 6-deoxy-N-acetyl-alpha-D-galactosamine 1-phosphate
show the reaction diagram
-
37% phosphorylation yield
-
-
?
ATP + 6-deoxy-N-acetyl-D-glucosamine
ADP + 6-deoxy-N-acetyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
75% phosphorylation yield
-
-
?
ATP + 6-methyl-N-acetyl-D-mannosamine
ADP + 6-methyl-N-acetyl-alpha-D-mannosamine 1-phosphate
show the reaction diagram
ATP + D-galactopyranose
ADP + alpha-D-galactopyranose 1-phosphate
show the reaction diagram
-
below 5% phosphorylation yield
-
-
?
ATP + D-galactosamine
ADP + D-galactosamine 1-phosphate
show the reaction diagram
ATP + D-galactose
ADP + D-galactose 1-phosphate
show the reaction diagram
ATP + D-glucosamine
ADP + D-glucosamine 1-phosphate
show the reaction diagram
-
-
-
?
ATP + D-glucose
ADP + D-glucose 1-phosphate
show the reaction diagram
low activity
-
-
?
ATP + D-mannosamine
ADP + alpha-D-mannosamine 1-phosphate
show the reaction diagram
-
-
-
?
ATP + D-mannose
?
show the reaction diagram
ATP + D-mannose
ADP + alpha-D-mannose 1-phosphate
show the reaction diagram
ATP + D-talose
?
show the reaction diagram
phosphorylation at 3% of the activity with N-acetylglucosamine
-
-
?
ATP + mannose
ADP + mannose 1-phosphate
show the reaction diagram
-
-
-
?
ATP + N-acetyl-alpha-D-galactosamine
ADP + N-acetyl-alpha-D-galactosamine 1-phosphate
show the reaction diagram
ATP + N-acetyl-alpha-D-glucosamine
ADP + N-acetyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
ATP + N-acetyl-D-galactosamine
ADP + N-acetyl-alpha-D-galactosamine 1-phosphate
show the reaction diagram
ATP + N-acetyl-D-glucosamine
ADP + N-acetyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
ATP + N-acetyl-D-hexosamine
ADP + N-acetyl-alpha-D-hexosamine 1-phosphate
show the reaction diagram
ATP + N-acetyl-D-mannosamine
?
show the reaction diagram
phosphorylation at 15% of the activity with N-acetylglucosamine
-
-
?
ATP + N-acetyl-D-mannosamine
ADP + ?
show the reaction diagram
-
-
-
?
ATP + N-acetyl-D-mannosamine
ADP + N-acetyl-alpha-D-mannosamine 1-phosphate
show the reaction diagram
low activity
-
-
?
ATP + N-azidoacetyl-D-galactosamine
ADP + N-azidoacetyl-alpha-D-galactosamine 1-phosphate
show the reaction diagram
-
65% phosphorylation yield
-
-
?
ATP + N-azidoacetyl-D-glucosamine
ADP + N-azidoacetyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
87% phosphorylation yield
-
-
?
ATP + N-benzoyl-alpha-D-galactosamine
ADP + N-benzoyl-alpha-D-galactosamine 1-phosphate
show the reaction diagram
ATP + N-butanoyl-D-galactosamine
ADP + N-butanoyl-alpha-D-galactosamine 1-phosphate
show the reaction diagram
ATP + N-butanoyl-D-glucosamine
ADP + N-butanoyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
ATP + N-phenylacetyl-D-galactosamine
ADP + N-phenylacetyl-alpha-D-galactosamine 1-phosphate
show the reaction diagram
-
77% phosphorylation yield
-
-
?
ATP + N-phenylacetyl-D-glucosamine
ADP + N-phenylacetyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
88% phosphorylation yield
-
-
?
ATP + N-propionyl-D-galactosamine
ADP + N-propionyl-alpha-D-galactosamine 1-phosphate
show the reaction diagram
-
85% phosphorylation yield
-
-
?
ATP + N-propionyl-D-glucosamine
ADP + N-propionyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
86% phosphorylation yield
-
-
?
ATP + N-trifluoroacetyl-alpha-D-galactosamine
ADP + N-trifluoroacetyl-alpha-D-galactosamine 1-phosphate
show the reaction diagram
-
0.75 microM, 21.8% conversion
-
-
?
ATP + N-trifluoroacetyl-D-glucosamine
ADP + N-trifluoroacetyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
-
recombinant enzyme
-
-
?
GTP + N-acetyl-alpha-D-glucosamine
GDP + N-acetyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
GTP shows about 45% of the activity with ATP
-
-
?
ITP + N-acetyl-alpha-D-glucosamine
IDP + N-acetyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
ITP shows about 30% of the activity with ATP
-
-
?
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
ATP + N-acetyl-alpha-D-galactosamine
ADP + N-acetyl-alpha-D-galactosamine 1-phosphate
show the reaction diagram
ATP + N-acetyl-alpha-D-glucosamine
ADP + N-acetyl-alpha-D-glucosamine 1-phosphate
show the reaction diagram
ATP + N-acetyl-D-hexosamine
ADP + N-acetyl-alpha-D-hexosamine 1-phosphate
show the reaction diagram
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ca2+
1 mM, 1% relative activity compared to Mg2+
Co2+
1 mM, 21% relative activity compared to Mg2+; 21% of the activity with Mg2+
MgCl2
-
10 mM are included in the assay medium
Mn2+
1 mM, 7% relative activity compared to Mg2+
Ni2+
1 mM, 4% relative activity compared to Mg2+
Zn2+
1 mM, 8% relative activity compared to Mg2+
additional information
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
isopropyl-1-thio-beta-D-galactoside
induction of protein expression at a final concentration of 0.5 mM
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.005 - 0.9
ATP
0.065 - 0.09
N-acetyl-alpha-D-galactosamine
0.06 - 0.118
N-acetyl-alpha-D-glucosamine
0.35 - 3.14
N-acetyl-D-glucosamine
additional information
additional information
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summary of ITC analysis of wild-type NahK and mutants
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.12 - 16.3
ATP
0.46 - 6.08
N-acetyl-alpha-D-galactosamine
0.95 - 1.21
N-acetyl-alpha-D-glucosamine
0.11 - 13.6
N-acetyl-D-glucosamine
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kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
4.8 - 11.8
ATP
5.1 - 7.1
N-acetyl-alpha-D-galactosamine
15.8 - 18.3
N-acetyl-alpha-D-glucosamine
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.001
D-glucose, substrate donor specificity
0.002
D-mannosamine, substrate donor specificity
0.004
D-glucosamine, substrate donor specificity
0.007
2-deoxyglucose, substrate donor specificity
0.024
D-mannose, substrate donor specificity
0.049
D-talose, substrate donor specificity
0.22
N-acetyl-D-mannosamine, substrate donor specificity
0.48
ITP, substrate donor specificity
0.65
GTP, substrate donor specificity
0.89
N-acetyl-D-galactosamine, substrate donor specificity
1.48
ATP, substrate donor specificity
1.49
N-acetyl-D-glucosamine, substrate donor specificity
additional information
Incubation of 4 microM LnpB protein with 10 mM N-acetyl-alpha-D-glucosamine 6-sulfate in 0.1 M MOPS buffer (pH 7.5) at 30°C for 30 min shows that the protein have no hydrolytic activity
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TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
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SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
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PDB
SCOP
CATH
ORGANISM
UNIPROT
Bifidobacterium longum subsp. infantis (strain ATCC 15697 / DSM 20088 / JCM 1222 / NCTC 11817 / S12)
Bifidobacterium longum subsp. infantis (strain ATCC 15697 / DSM 20088 / JCM 1222 / NCTC 11817 / S12)
Bifidobacterium longum subsp. longum (strain ATCC 15707 / DSM 20219 / JCM 1217 / NCTC 11818 / E194b)
Bifidobacterium longum subsp. longum (strain ATCC 15707 / DSM 20219 / JCM 1217 / NCTC 11818 / E194b)
Bifidobacterium longum subsp. longum (strain ATCC 15707 / DSM 20219 / JCM 1217 / NCTC 11818 / E194b)
Bifidobacterium longum subsp. longum (strain ATCC 15707 / DSM 20219 / JCM 1217 / NCTC 11818 / E194b)
Bifidobacterium longum subsp. longum (strain ATCC 15707 / DSM 20219 / JCM 1217 / NCTC 11818 / E194b)
Bifidobacterium longum subsp. longum (strain ATCC 15707 / DSM 20219 / JCM 1217 / NCTC 11818 / E194b)
Bifidobacterium longum subsp. longum (strain ATCC 15707 / DSM 20219 / JCM 1217 / NCTC 11818 / E194b)
Bifidobacterium longum subsp. longum (strain ATCC 15707 / DSM 20219 / JCM 1217 / NCTC 11818 / E194b)
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MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
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SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
additional information
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Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified recombinant His-tagged enzyme in complex with substrates GalNAc, GlcNAc-1P, GlcNAc/AMPPNP (a nonhydrolyzable ATP analogue) and GlcNAc-1P/ADP, hanging drop vapour diffusion method, pre-incubation of SeMet-substituted/wild-type NahK with GlcNAc (20fold excess), GalNAc (20fold excess) or GlcNAc-1P (10fold excess), and mixing of 20 mg/ml protein in 50 mM HEPES, pH 7.5, with a crystallization solution containing 20% w/v PEG 3350, 0.1 M Bis-Tris, pH 5.5, crystals of NahK-GlcNAc-AMPPNP are obtained by soaking NahK-GlcNAc crystals with 10 mM AMPPNP and 20 mM MgCl2 for 15 min, or the crystals of NahK-GlcNAc-1P-ADP by soaking NahK-GlcNAc-1P crystals with 5 mM ADP and 10 mM MgCl2 for 30 min, crystallization time is 10-14 days, 20°C, X-ray diffraction structure determination and analysis at 1.72-2.15 A resolution, molecular replacement method, modelling
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purified recombinant wild-type enzyme in apoform, or SeMet-labeled NahK complexed with ATP, sitting drop vapor diffusion method, mixing of 500 nl protein solution containing 25 mg/mL NahK and with or without 10 mM ATP with an equal volume of reservoir solution containing 0.2 M magnesium acetate, 0.1 M sodium cacodylate, pH 6.5, and 20% PEG 8000, for ADP complex crystals, 10 mM ADP is used instead of ATP, 20°C, X-ray diffraction structure determination and analysis at 1.80-2.05 A resolution
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purified recombinant His-tagged enzyme in complex with substrates GlcNAc, and GlcNAc/AMPPNP (a nonhydrolyzable ATP analogue), hanging drop vapour diffusion method, mixing of 20 mg/ml protein in 50 mM HEPES, pH 7.5, with crystallization solution containing 1.5 M Li2SO4, 0.1 M sodium acetate, pH 5.5, 2 days, crystals of the NahKATCC15697-GlcNAc-AMPPNP complex are obtained by soaking NahK-GlcNAc complex crystals with 5 mM AMPPNP and 10 mM MgCl2 for 1.5 h, 20°C, X-ray diffraction structure determination and analysis at 1.47-1.90 A resolution, molecular replacement method, modelling
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 9.5
stability at 30°C; stable
677700
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50
pH 8.5, 30 min, 50% loss of activity
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GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
loses half of enzyme activity during incubation for 30 min at 50°C in 0.1 M Tris buffer (pH 8.5)
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
; The transformant is cultivated and protein expression is induced by the addition of isopropyl-1-thio-beta-D-galactoside. The cells are harvested by centrifugation and sonicated. Cell debris is removed by centrifugation and enzyme is purified on an Ni-nitrilotriacetic acid agarose gel.
recombinant C-terminally His6-tagged wild-type and SeMet-labeled NahK residues 1-359 enzymes from Escherichia coli by nickel affinity chromatography and gel filtration
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recombinant N-terminally His6-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and gel filtration
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Escherichia coli strains select96 and BL21(DE3) are used as hosts for cloning and expression.; expression in Escherichia coli
expression in Escherichia coli
gene lnpB, recombinant expression of C-terminally His6-tagged NahK residues 1-359 in Escherichia coli strain BL21 CodonPlus (DE3)-RIL, expression of the (SeMet)-labeled protein in Escherichia coli strain B834 (DE3)
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gene lnpB, recombinant expression of N-terminally His6-tagged enzyme in Escherichia coli strain BL21(DE3)
recombinant expression in Escherichia coli strain BL21(DE3)
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D208 A
-
site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to the wild-type
D208A
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site-directed mutagenesis, the mutant shows altered activity and kinetics compared to the wild-type enzyme
D208A/K210A
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site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to the wild-type
D208A/K210L
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site-directed mutagenesis, the mutant shows highly reduced activity and altered substrate specificity compared to the wild-type
D208L
-
site-directed mutagenesis, the mutant shows highly reduced activity and altered substrate specificity compared to the wild-type
D228A
-
site-directed mutagenesis, the mutant shows altered activity and kinetics compared to the wild-type enzyme
I146E
-
site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to the wild-type
I146E/T231E
-
site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to the wild-type
K210E
-
site-directed mutagenesis, the mutant shows reduced highly activity and altered substrate specificity compared to the wild-type
K210L
-
site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to the wild-type
N213A
-
site-directed mutagenesis, the mutant shows altered activity and kinetics compared to the wild-type enzyme
Q48A
-
site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to the wild-type
R306A
-
site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to the wild-type
R306A/Y317A
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site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to the wild-type
R306E
-
site-directed mutagenesis, the mutant shows highly reduced activity but unaltered substrate specificity compared to the wild-type
R306E/Y317F
-
site-directed mutagenesis, the mutant shows highly reduced activity but unaltered substrate specificity compared to the wild-type
T231 E
-
site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to the wild-type
Y317A
-
site-directed mutagenesis, the mutant shows similar activity but slightly altered substrate specificity compared to the wild-type
Y317F
-
site-directed mutagenesis, the mutant shows similar activity but slightly altered substrate specificity compared to the wild-type
D208A
-
site-directed mutagenesis, the mutant shows altered activity and kinetics compared to the wild-type enzyme
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D228A
-
site-directed mutagenesis, the mutant shows altered activity and kinetics compared to the wild-type enzyme
-
K210A
-
site-directed mutagenesis, the mutant shows altered activity and kinetics compared to the wild-type enzyme
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N213A
-
site-directed mutagenesis, the mutant shows altered activity and kinetics compared to the wild-type enzyme
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Q48A
-
site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to the wild-type
-
R306A
-
site-directed mutagenesis, the mutant shows reduced activity and altered substrate specificity compared to the wild-type
-
R306E
-
site-directed mutagenesis, the mutant shows highly reduced activity but unaltered substrate specificity compared to the wild-type
-
Y317A
-
site-directed mutagenesis, the mutant shows similar activity but slightly altered substrate specificity compared to the wild-type
-
Y317F
-
site-directed mutagenesis, the mutant shows similar activity but slightly altered substrate specificity compared to the wild-type
-
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
synthesis
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LINKS TO OTHER DATABASES (specific for EC-Number 2.7.1.162)
ExplorEnz
ExPASy
KEGG
MetaCyc
SABIO-RK
NCBI: PubMed, Protein, Nucleotide, Structure, Genome, OMIM
IUBMB Enzyme Nomenclature
PROSITE Database of protein families and domains
InterPro (database of protein families, domains and functional sites)