Information on EC 2.7.1.52 - fucokinase

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

EC NUMBER
COMMENTARY hide
2.7.1.52
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RECOMMENDED NAME
GeneOntology No.
fucokinase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + L-fucose = ADP + beta-L-fucose 1-phosphate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phospho group transfer
-
-
-
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Amino sugar and nucleotide sugar metabolism
-
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Fructose and mannose metabolism
-
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GDP-L-fucose biosynthesis II (from L-fucose)
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Metabolic pathways
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SYSTEMATIC NAME
IUBMB Comments
ATP:beta-L-fucose 1-phosphotransferase
Requires a divalent cation for activity, with Mg2+ and Fe2+ giving rise to the highest enzyme activity. Forms part of a salvage pathway for reutilization of L-fucose. Can also phosphorylate D-arabinose, but more slowly.
CAS REGISTRY NUMBER
COMMENTARY hide
37278-00-5
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
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Uniprot
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
no activity in Saccharomyces cerevisiae
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
metabolism
physiological function
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ATP + 6-deoxy-L-galactose
ADP + 6-deoxy-L-galactose 1-phosphate
show the reaction diagram
ATP + D-arabinose
?
show the reaction diagram
ATP + D-glucose
?
show the reaction diagram
ATP + D-ribose
?
show the reaction diagram
-
87% of the activity with L-fucose
-
-
?
ATP + L-arabinose
?
show the reaction diagram
-
31% of the activity with L-fucose
-
-
?
ATP + L-fucose
ADP + beta-L-fucose 1-phosphate
show the reaction diagram
ATP + L-rhamnose
?
show the reaction diagram
-
41% of the activity with L-fucose
-
-
?
CTP + 6-deoxy-L-galactose
CDP + 6-deoxy-L-galactose 1-phosphate
show the reaction diagram
GTP + 6-deoxy-L-galactose
GDP + 6-deoxy-L-galactose 1-phosphate
show the reaction diagram
ITP + 6-deoxy-L-galactose
IDP + 6-deoxy-L-galactose 1-phosphate
show the reaction diagram
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2.5% of the activity with L-fucose
-
-
?
L-fucose + ATP
beta-L-fucose 1-phosphate + ADP
show the reaction diagram
UTP + 6-deoxy-L-galactose
UDP + 6-deoxy-L-galactose 1-phosphate
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
ATP + 6-deoxy-L-galactose
ADP + 6-deoxy-L-galactose 1-phosphate
show the reaction diagram
ATP + L-fucose
ADP + beta-L-fucose 1-phosphate
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ca2+
-
absolute requirement for a divalent cation, 85% of the activation with Mg2+
additional information
-
not influenced by Ca2+, Cu2+, K+, Hg2+, Ni2+ and Zn2+
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1,1,1-trichlorethyl 6-deoxy-alpha-L-galactopyranoside
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10 mM, 87% inhibition
2-fluoro-L-fucose
-
-
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4,6-dideoxy-4-azido-alpha-L-glucopyranose
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10 mM, 9% inhibition
4,6-dideoxy-4-fluoro-L-gluco-pyranose
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10 mM, 30% inhibition
4,6-dideoxy-4-iodo-L-glucopyranose
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10 mM, 75% inhibition
4,6-dideoxy-L-xylo-hexopyranose
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10 mM, 74% inhibition
ADP
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4.9 mM, 96% inhibition, competitive with ATP
AMP
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5.0 mM, 14% inhibition in presence of ATP
CDP
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4.9 mM, 16% inhibition in presence of ATP
Co2+
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complete inhibition at 20 mM
CTP
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4.9 mM, 30% inhibition in presence of ATP
D-arabinose
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0.5 mM, 20% inhibition. 10 mM, 84% inhibition
Fe2+
-
complete inhibition at 20 mM
GDP-L-fucose
GTP
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0.1 mM, 59% inhibition in presence of ATP
L-fucose
L-galactose
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weak
methyl 3,6-dideoxy-alpha-L-xylo-hexopyranoside
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10 mM, 17% inhibition
methyl 4,6-dideoxy-4-iodo-alpha-L-glucopyranoside
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10 mM, 54% inhibition
methyl 4,6-dideoxy-alpha-L-xylo-hexopyranoside
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10 mM, 9% inhibition
methyl 6-deoxy-2-O-dodecanoyl-alpha-L-galactopyranoside
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10 mM, 37% inhibition
methyl 6-deoxy-alpha-L-galactopyranoside
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10 mM, 89% inhibition
octyl 6-deoxy-alpha-L-galactopyranoside
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10 mM, 40% inhibition
TTP
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4.8 mM, 45% inhibition in presence of ATP
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
GDP-alpha-D-mannose
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stimulation
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.29 - 0.8
ATP
0.017 - 1.59
L-fucose
1
MgATP2-
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pH 7.4, 37°C
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.22 - 0.38
ATP
0.34 - 23.35
L-fucose
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
5.5
1,1,1-trichlorethyl 6-deoxy-alpha-L-galactopyranoside
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pH 7.4, 37°C
5
4,6-dideoxy-4-iodo-L-glucopyranose
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pH 7.4, 37°C
0.5
4,6-dideoxy-L-xylo-hexopyranose
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pH 7.4, 37°C
0.01
GDP-L-fucose
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-
1.1
methyl 6-deoxy-alpha-L-galactopyranoside
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pH 7.4, 37°C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.003275
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-
0.0091
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radiometric assay
0.6
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-
1.273
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-
82.9
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pH 7.5, 35°C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.6
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8.3
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two optima: pH 6.5 and pH 8.3
10.5
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optimum for fucokinase activity
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 10
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more than 50% activity between pH 6.0 and 10.0
7 - 9.5
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pH 7: about 50% of maximal activity, pH 9.5: about 60% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
40
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optimum for fucokinase activity
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25 - 50
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more than 50% activity between 25 and 50°C
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.9
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calculated
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
additional information
-
expression in all tissues examined
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
78000
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4 * 78000, SDS-PAGE
110000
111000
short splicing variant
116350
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x * 116350, calculated, x * 120000, SDS-PAGE
118800
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gel filtration
120000
309200
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gel filtration
315300
analytical ultracentrifugation
440000
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gel filtration
494000
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homotrimer
3 * 105000, recombinant enzyme, calculated from amino acid sequence
monomer
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1 * 118800
tetramer
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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peptide sequence does not contain a secretory signal or transmembrane domain
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
hanging drop vapor diffusion method, using 20% (w/v) PEG 3350 as precipitant, 200 mM trisodium citrate pH 8.3
purified apo-enzyme, mixing of 10 mg/ml protein in 10 mM Tris-HCl pH 7.5 with 9% w/v PEG 6K, 5% v/v ethylene glycol, 5% v/v glycerol, 25 mM HEPES pH 7.5, 2% tert-butanol, 1 mM MgCl2, 1mM MnCl2, 24°C, 20-30 days, X-ray diffraction structure determination and analysis at 2.6 A resolution
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3 - 12
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After 30 min at pH 3,0, 5.0, 7.0, 8.0, 10.0, and 12.0, the enzyme shows about 25%, 50%, 90%, 100%, 75%, and 35% activity, respectively
738343
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4 - 37
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the enzyme is stable for 2 h at 4 and 12°C. At 20°C, the enzyme shows 80% activity after 1 h and less than 20% activity after 2 h incubation. At 28°C, the enzyme is stable for 30 min, while at 37°C, the enzyme activity drops to about 37% after 30 min
50
-
loss of 99% of activity
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
EDTA stabilizes during purification
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glycerol, sucrose, DTT or 2-mercaptoethanol stabilizes
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instability of the enzyme occurs through chemical reduction in disulfide bonds of FKP protein, resulting in degradation
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ovalbumin does not stabilize purified enzyme
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ovalbumin stabilizes
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substrates do not stabilize
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unstable in the presence of KCl
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-16°C, about 30% loss of activity within 24 h and about 60% loss of activity within 72 h
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-16°C, in the presence of stabilizing ovalbumin, about 30% loss of activity within 48 h
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-20°C, 50% glycerol, 1 mM PMSF, 0.2 M NaCl, 50 mM Tris-HCl buffer, pH 7.5, stable for 1 month
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-20°C, in the presence of 2-mercaptoethanol and glycerol, quite stable
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-80°C, crude extracts, at least 2 years, without significant loss of activity
2-4°C, most stable in the presence of glycerol and a sulfhydryl reagent
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4°C, 1 day, loss of 50% of GDP-L-fucose diphosphorylase activity
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4°C, unstable in saline solution
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frozen, in the presence of glycerol or sucrose and a sulfhydryl reagent, up to 1 week
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frozen, rapid loss of activity, glycerol or sucrose and DTT or 2-mercaptoethanol stabilize, no stabilization by Mg2+, EDTA, KCl or ovalbumin
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frozen, unstable in saline solution
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Ni-affinity column chromatography and Superdex G200 gel filtration
nickel affinity column chromatography
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partial
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recombinant maltose-binding protein fusion protein from Escherichia coli strain BL21(DE3) by amylose affinity chromatography, cleavage of the tag by Genease I protease, followed by guanosine affinity chromatography and ultrafiltration
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recombinant protein with His-tag and fused to thioredoxin, partial purification
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli BL21(DE3) cells
expression as fusion protein with maltose-binding protein in Escherichia coli strain BL21 (DE3)
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expression in Escherichia coli with chaperones GroEL, GroES
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expression of full-length protein and a deletion mutant lacking the first 363 amino acids of the N-terminus are expressed in Escherichia coli BL21 cells. Both proteins display fucokinase activity. Identification of amino acid sequence
functional expression of HA-tagged FKP in Saccharomyces cerevisiae strain W303-1A, the biosynthesis of GDP-D-Ara in the recombinant Saccharomyces cerevisiae strain harboring the FKP gene can occur through a mechanism akin to that of GDP-L-Fuc via the salvage pathway. Expression of a loss-of-function FKP mutant in yeast
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two different transcripts of 3057 and 3270 base pairs expressed in COS-7 cells, only the longer splice variant is enzymatically active; two different transcripts of 3057 and 3270 base pairs expressed in COS-7 cells, only the longer splice variant is enzymatically active
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
the enzyme transcript level is up-regulated 4.3fold by nitrogen exhaustion when Mortierella alpina starts to accumulate lipid
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
G133A
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mutation abolishes GDP-L-fucose diphosphorylase activity, hardly affects fucokinase activity
G830A
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mutation reduces fucokinase activity by more than 90%, while preserving the GDP-L-fucose diphosphorylase activity
additional information
deletion mutant lacking the first 363 amino acids of the N-terminus expressed in Escherichia coli BL21 cells display fucokinase activity
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
synthesis
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the enzyme is a valuable biochemical tool to prepare activated L-fucose derivatives for fucosylation reactions, enzyme is valuable for the formation of radiolabeled fucose 1-phosphate