Information on EC 2.7.1.30 - glycerol kinase

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

EC NUMBER
COMMENTARY hide
2.7.1.30
-
RECOMMENDED NAME
GeneOntology No.
glycerol kinase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + glycerol = ADP + sn-glycerol 3-phosphate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phospho group transfer
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
degradation of sugar alcohols
-
-
glycerol degradation I
-
-
Glycerolipid metabolism
-
-
Metabolic pathways
-
-
SYSTEMATIC NAME
IUBMB Comments
ATP:glycerol 3-phosphotransferase
Glycerone and L-glyceraldehyde can act as acceptors; UTP (and, in the case of the yeast enzyme, ITP and GTP) can act as donors.
CAS REGISTRY NUMBER
COMMENTARY hide
9030-66-4
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
oat
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
queen bumblebee
-
-
Manually annotated by BRENDA team
Candida mycoderma
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
glycerol kinase from Cellulomonas sp. is used to develop a bisensor based on flow calorimetry for quantitative analysis of glycerol during bioconversion process. Thermomeric glycerol sensor employs flow calorimeter equipped with a column packed with immobilized glycerol kinase. The glycerol kinase from Escherichia coli is able to detect glycerol in samples, but the linearity range and sensitibity are rather low and the calibration line does not pass through the origin of calibration dependence. The enzyme from Cellulomonas sp. provides a similar trend but significantly better results. The immobilized enzyme stability is excellent. The immobilized enzyme column is stored at 4C. It can do 1 month of total operation time at 30C with no significant loss of sensitivity. No interference with 1,3-propanediol and fermentation medium is observable.
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
pigeon
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
there are five glycerol kinase-like genes in Drosophila melanogaster. The locus CG18374, located at 61B2, is the closest homolog to the X-linked glycosyl kinase gene in Homo sapiens, sharing 53% identity. Another close homolog CG7995 (located at 62B1) shares 48% homology to the human glycerol kinase. Three other loci CG1271 at 63A3, CG1216 at 61B2, and CG8298 at 48D5 do not share the same degree of homology. Most key functional residues are missing in CG1216 suggesting it has no glycerol kinase activity.; there are five glycerol kinase-like genes in Drosophila melanogaster.
-
-
Manually annotated by BRENDA team
Escherichia coli C.Lin 43
strain C.Lin 43
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
Mycobacterium butyricum
-
-
-
Manually annotated by BRENDA team
607
-
-
Manually annotated by BRENDA team
607
-
-
Manually annotated by BRENDA team
rainbow smelt
-
-
Manually annotated by BRENDA team
kidney bean
-
-
Manually annotated by BRENDA team
pea
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
trout
-
-
-
Manually annotated by BRENDA team
-
UniProt
Manually annotated by BRENDA team
-
Uniprot
Manually annotated by BRENDA team
-
UniProt
Manually annotated by BRENDA team
-
UniProt
Manually annotated by BRENDA team
-
UniProt
Manually annotated by BRENDA team
broad bean
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
metabolism
-
the enzyme plays an essential role in central and lipid 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
ADP + sn-glycerol 3-phosphate
ATP + glycerol
show the reaction diagram
ATP + 1,3-propanediol
ADP + ?
show the reaction diagram
Candida mycoderma
-
weak
-
-
?
ATP + 1-deoxy-sn-glycerol
ADP + ?
show the reaction diagram
Candida mycoderma
-
-
-
-
?
ATP + 2-deoxyglycerol
ADP + ?
show the reaction diagram
Candida mycoderma
-
-
-
-
?
ATP + 2-mercaptoethanol
ADP + ?
show the reaction diagram
Candida mycoderma
-
-
-
-
?
ATP + 2-methylglycerol
ADP + ?
show the reaction diagram
Candida mycoderma
-
-
-
-
?
ATP + aminopropanediol
ADP + ?
show the reaction diagram
Candida mycoderma
-
R- and S-
-
-
?
ATP + D-glyceraldehyde
ADP + D-glyceraldehyde 3-phosphate
show the reaction diagram
-
-
-
-
?
ATP + dihydroxyacetone
ADP + dihydroxyacetone phosphate
show the reaction diagram
ATP + dihydroxypropyl dichloroacetate
ADP + ?
show the reaction diagram
-
glycerol analogue
-
-
?
ATP + glyceric acid
ADP + ?
show the reaction diagram
-
-
-
-
?
ATP + glycerol
?
show the reaction diagram
ATP + glycerol
ADP + sn-glycerol 3-phosphate
show the reaction diagram
ATP + L-glyceraldehyde
ADP + L-glyceraldehyde 3-phosphate
show the reaction diagram
ATP + mercaptopropanediol
1-mercaptopropanediol 1-phosphate + ADP
show the reaction diagram
ATP + monoacetin
ADP + ?
show the reaction diagram
-
glycerol analogue
-
-
?
ATP + monobutyrin
ADP + ?
show the reaction diagram
-
glycerol analogue
-
-
?
ATP + monothioglycerol
ADP + ?
show the reaction diagram
-
-
-
-
?
CTP + glycerol
CDP + glycerol 3-phosphate
show the reaction diagram
glycerol + ATP
sn-glycerol 3-phosphate + ADP
show the reaction diagram
GTP + glycerol
GDP + glycerol 3-phosphate
show the reaction diagram
ITP + glycerol
IDP + glycerol 3-phosphate
show the reaction diagram
TTP + glycerol
TDP + glycerol 3-phosphate
show the reaction diagram
-
-
-
-
?
UTP + glycerol
UDP + glycerol 3-phosphate
show the reaction diagram
XTP + glycerol
XDP + glycerol 3-phosphate
show the reaction diagram
Candida mycoderma
-
-
-
-
?
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 + glycerol
?
show the reaction diagram
ATP + glycerol
ADP + sn-glycerol 3-phosphate
show the reaction diagram
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
(NH4)2SO4
-
activates
Co2+
-
best effect, may be replaced by Zn2+
Zn2+
-
can substitute for Co2+
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1,2-propanediol
-
-
1-Pentanol
-
-
3-Chloro-1,2-propanediol
-
-
3-Deoxy-sn-glycerol
Candida mycoderma
-
-
ATP
-
substrate inhibition
Butane-1,3-diol
Candida mycoderma
-
D- and L-configuration
CrATP
Candida mycoderma
-
coordination complex of Cr3+ and ATP, dead-end inhibitor
cytosolic subunit of the glucose-specific phosphotransferase system
-
allosteric inhibition
-
D-fructose 1,6-bisphosphate
-
-
D-fructose-1,6-bisphosphate
-
allosteric inhibition
DTNB
-
inactivation reversed by dithiothreitol
erythritol
Candida mycoderma
-
-
ethanediol
ethanol
Candida mycoderma
-
-
fructose 1,6-bisphosphate
fructose 1,6-diphosphate
glucose
glucose 6-phosphate
glucose-specific phosphocarrier protein of the phosphoenolpyruvate:glucose phosphotransferase system
-
-
-
glucose-specific phosphocarrier protein of the phosphoenolpyruvate:glucose phosphotransferase system (IIA(Glc))
-
allosteric inhibitor
-
glycerol
glycerol 3-phosphate
iodoacetamide
iodoacetate
-
-
L(+)-Butane-2,3-diol
Candida mycoderma
-
-
L-alpha-glycerophosphate
-
no inhibition up to 3 mM in presence of 0.1 M glycerol
L-Threitol
Candida mycoderma
-
-
N-ethylmaleimide
p-chloromercuribenzoate
p-Hydroxymercuriphenylsulfonate
-
-
phosphocarrier protein IIAGlc
-
the unphosphorylated form of the phosphocarrier protein IIAGlc is an allosteric inhibitor of Escherichia coli glycerol kinase
-
Procion Blue MX-3G
-
5 mM, inactivates after a period of increased activity
propan-1-ol
propan-2-ol
Salyrganic acid
Candida mycoderma
-
-
additional information
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1,3-butanediol
-
activates
1,3-Propanediol
-
activates
1,4-Butanediol
-
activates
1-butanol
-
activates
1-Chloro-2-propanol
-
activates
1-propanol
2,3-Butanediol
-
activates
2-butanol
-
activates
2-Chloroethanol
-
activates
2-methyl-1-propanol
-
activates
2-Methyl-2-propanol
-
activates
2-Pentanol
-
activates
2-propanol
3-methyl-1-butanol
-
activates
Cyclohexanol
-
activates
Ethanediol methyl ether
-
activates
ethanol
-
activates
methanol
-
activates
rosiglitazone
-
rosiglitazone treatment markebly increases glycerol kinase mRNA expression in both the mesenteric and epididymal adipose tissues (1085.9% and 523.8% of the abundance in Long-Evans Tokushima Otsuka rats, respectively). The magnitude of glycerol kinase induced by rosiglitazone is significantly greater in the mesenteric fat than in the epididymal fat.
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
45
1-deoxy-sn-glycerol
Candida mycoderma
-
-
88
2-deoxyglycerol
Candida mycoderma
-
-
0.046
2-mercaptoethanol
Candida mycoderma
-
25C, pH 8.2
5.7
2-methylglycerol
Candida mycoderma
-
-
0.24 - 0.69
ADP
0.006 - 3.29
ATP
0.515
CTP
-
-
0.14
dichloro-monoacetin
-
pH 7.35, 37C
0.5 - 100
dihydroxyacetone
0.15
glyceric acid
0.01 - 2
glycerol
12.56
glycerol-3-phosphate
-
pH 7.0
0.145
GTP
-
-
3 - 42
L-Glyceraldehyde
3.83
L-glycerol-3-phosphate
-
25C, pH 8.0
0.009 - 0.01
MgATP2-
0.169
Monoacetin
-
pH 7.35, 37C
0.491
Monobutyrin
-
pH 7.35, 37C
4.9
monothioglycerol
1.09 - 1.37
sn-glycerol 3-phosphate
0.62
UTP
-
-
additional information
additional information
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1670
glycerol
Candida mycoderma
-
-
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
250
propan-1-ol
Candida mycoderma
-
-
1500
propan-2-ol
Candida mycoderma
-
-
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.001 - 0.01
glucose-specific phosphocarrier protein of the phosphoenolpyruvate:glucose phosphotransferase system (IIA(Glc))
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.054
-
recombinant enzyme, using 1% methanol as carbon sources in the medium, at pH 9.8 and 30C
0.13
-
recombinant enzyme, using 1% glycerol and 1% methanol as carbon sources in the medium, at pH 9.8 and 30C
2
-
at pH 7.0 and 70C
15.5
for glycerol
25.2
-
-
41.2
-
pH 7.0, 25C
101
Candida mycoderma
-
25C
177
-
pH 7.5, 25C
additional information
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.8 - 10.5
-
pH 6.8: about 30% of maximum activity, pH 10.5: about 80% of maximum activity
7 - 9.5
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37
-
assay at
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 80
-
30C: about 50% of maximum activity, 80C: about 65% of maximum activity
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
-
lung cell line
Manually annotated by BRENDA team
-
There is no difference in glycerol mRNA level between control 3T3-L1 adipocytes and 3T3-L1 Aqp7-RNAi transfected adipocytes, whereas a 4fold enzymatic activation of glycerol kinase is observable in Aqp7 knockout adipocytes (activity assay: 50 mM TrisHCl, pH 7.2, 5 mM ATP, 10 mM MgCl2, 100 mM KCl, 2.5 mM DTT, 4 mM glycerol, 500 microM 3H-glycerol, for 90 min at 37C).
Manually annotated by BRENDA team
-
SS6-2 cell, stem cells from fat with myotube characteristics
Manually annotated by BRENDA team
-
Aqp7-/- knockout mice have significantly lower plasma glycerol level under 12 h fasting conditions.
Manually annotated by BRENDA team
-
newborn, expresses only isoform GK-EX18
Manually annotated by BRENDA team
-
from patients with enzyme deficiency and normal individuals
Manually annotated by BRENDA team
-
kidney cell line
Manually annotated by BRENDA team
-
kidney cell line, expresses only isoform GK+EX18
Manually annotated by BRENDA team
-
adult hepatoblastoma cell line
Manually annotated by BRENDA team
-
primary culture of
Manually annotated by BRENDA team
-
4th instar larvae
Manually annotated by BRENDA team
Candida mycoderma
-
-
Manually annotated by BRENDA team
-
pancreatic islets and pancreas. The glycerol kinase mRNA level in the Aqp7-/- pancreatic islets is 655% that in Aqp7+/+ islets. The glycerol activity is increased in both total pancreases and islets isolated from Aqp7-/- mice compared with those isolated from Aqp+/+ mice (tested with cell homogenates, activity assay: 50 mM Tris-HCl, pH 7.2, 5 mM ATP, 10 mM MgCl2, 100 mM KCl, 2.5 mM dithiothreitol, 4 mM glycerol, 50 microM [14C]glycerol, for 3 h at 37C). The absence of AQP7 is associated with a moderate increase in the glycerol content of the total pancreas. There is a 2fold increase in the glycerol content of pancreatic islets isolated from Aqp7-/- mice compared with those from Aqp7+/+ mice.
Manually annotated by BRENDA team
-
expresses only isoform GK+EX18
Manually annotated by BRENDA team
-
fetal liver cell line
Manually annotated by BRENDA team
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
additional information
PDB
SCOP
CATH
ORGANISM
UNIPROT
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Rhizobium meliloti (strain 1021)
Staphylococcus aureus (strain COL)
Staphylococcus aureus (strain COL)
Thermococcus kodakarensis (strain ATCC BAA-918 / JCM 12380 / KOD1)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
54000 - 58000
-
gel filtration
113000
-
light scattering, dimer peak in the absence of fructose 1,6-bisphosphate, G230D mutant
120000
-
sucrose density gradient centrifugation
127000
-
light scattering, dimer peak in the presence of fructose 1,6-bisphosphate, G230D mutant
140000
-
gel filtration, zone sedimentation in sucrose density gradient
158000
-
dimeric native enzyme, gel filtration
177000
-
light scattering, dimer peak in the absence of fructose 1,6-bisphosphate, wild type enzyme
200000
-
gel filtration
210000 - 217000
-
equilibrium sedimentation
210000
-
gel filtration
220000
-
gel filtration
224000
-
tetrameric native enzyme, gel filtration
227000
-
light scattering, tetramer peak (about 2% of the principal peak) in the absence of fructose 1,6-bisphosphate, G230D mutant
230000
-
gel filtration
236000
-
gel filtration
251000
Candida mycoderma
-
diffusion and sedimentation data
280000 - 300000
-
sedimentation equilibrium light scattering method
391000
-
light scattering, tetramer peak (about 9% of the principal peak) in the absence of fructose 1,6-bisphosphate, wild type enzyme
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer or tetramer
-
2 or 4 * 60000, in solution, the enzyme exists in a dimer-tetramer equilibrium
homodimer
homotetramer
-
dimer-tetramer equilibrium in solution, tetramer in the crystal
monomer
-
1 * 53000
tetramer
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phosphoprotein
additional information
-
activity is enhanced phosphorylation of His232
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
crystals are obtained by the hanging-drop technique from a solution containing 29% polyethylene glycol 400, 0.1 M sodium acetate pH 4.5, 0.1 M calcium acetate and 10% glycerol. The crystals can grow in the presence of 33% PEG 400, which allows to mount the crystals and directyl flash-cool them. Repeated flash-annealing causes a significant decrease in the averaged mosaicity along with an increase in the overall peak counts of reflections and an enhanced signal-to-noise ratio. Individual reflection-profile analysis reveales a mostly dual domain structure, showing the minimization of one domain as a result of flash-annealing.
-
crystals of native and mutant enzyme with bound glycerol, hanging drop vapor diffusion method
-
in complex with glycerol, ADP and the allosteric effector enzyme IIAGlc
-
in complex with glycerol, in presence and absence of fructose 1,6-diphosphate, mechanism
-
of wild type and mutant A65T, both in complex with glycerol and ADP, and of mutant I474D, in complex with IIAGlc
-
the crystal structure of glycerol kinase mutant G230D is determined to 2.0 A resolution using a microfluidics based crystallization platform; using modified microfluidic scale-up diffraction device, crystals form after one week at ambient temperature unter crystallization conditions 0.3 M magnesium chloride, 0.1 M TrisHCl (pH 8.5), and 20% PEG 1500. Using vapor diffusion crystallization, crystals appear after one week at ambient temperature using the crystallization conditions 0.1 M magnesium chloride, 0.1 M TrisHCl (pH 8.5), and 10% PEG 1500. Glycerol kinase of the mutant G230D crystallied in space group P21 with two tetramers of 222 point symmetry in the asu. The average B factor for the overall structure of the G230D mutant is 21.2 A2.
-
crystals are grown at 20C by the sitting-drop vapour diffusion method. Native X-ray diffraction data are collected to 2.4 A resolution using synchrotron radiation at station BL44XU of SPring-8. The crystal belongs to the rhombohedral space group R3, with unit-cell parameters a = b = 217.48, c = 66.48 A. The protein is also cocrystallized with substrates and diffraction data are collected to 2.7 A resolution
using sitting-drop vapour-diffusion method at 293 K. Native Tk-glycerol kinase crystals appear after a few days using Wizard I solution No. 25 (0.1 M Tris pH 8.5, 0.2 M MgCl2, 30% PEG 400). Diffraction spots sufficient for structural determination at high resolution are not obtained when a crystal of Tk-glycerol kinase is mounted on a CryoLoop without cryoprotectant, diffraction patterns of the crystal are improved by using Paratone-N as cryoprotectant. Native X-ray diffraction data are collected to 2.4 A resolution using synchrotron radiation at station BL44XU of SPring-8. The crystals belong to the rhombohedral space group R3, with unit-cell parameters a = b = 217.48, c = 66.48 A. Assuming the presence of two molecules in the asymmetric unit, the VM value is 2.7 A3Da-1 and the solvent content is 54.1%.
-
sitting drop vapor diffusion method, using 30% (w/v) PEG 400 and 100 mM HEPES pH 7.0, at 20C
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 7
-
0C, half-life: 24 h
641295
6 - 9
-
30C, in presence of glycerol, completely stable for 30 min
641287
6 - 8
-
high stability
688443
6.7
Candida mycoderma
-
highest stability
641286
7.5
-
25C, no loss of activity after several h
641287
9.8
-
25C, half-life: 6.5 min
641287
additional information
-
glycerol affords considerable stabilization at the unfavorable pH values, glycerol kinases from microorganisms most stable at a neutral pH, glycerol kinases from higher organisms most stable in an acidic pH range
641287
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37
-
2 weeks, stable
68
-
30 min, pH 7.5, 50% activity
100
-
30 min, loss of 50% of activity
additional information
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
(NH4)2SO4 stabilizes
-
dialysis, against 0.01 M phosphate buffer, at 2C, pH 6.7, 50% loss of activity without glycerol, stable in presence of 0.01 M glycerol
Candida mycoderma
-
ethanediol stabilizes
Candida mycoderma
-
freezing without glycerol inactivates
-
glycerol affords considerable stabilization at the unfavorable pH values
-
glycerol, 0.01 M, EDTA, 0.001 M and 0.001 M 2-mercaptoethanol prevent inactivation during purification
-
normal and mutant enzyme stabilized against heat inactivation by glycerol, but not by fructose 1,6-bisphosphate
-
pigeon liver enzyme is sensitive to extreme dilution but can be stabilized by addition of 0.01% bovine serum albumin
-
the enzyme in crude extracts is stable to freezing and thawing, while more purified preparations are inactivated
OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
susceptible to inactivation by oxidation of sulfhydryl groups
-
641287
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, 50% glycerol, stable for more than 1 year
-
-20C, pH 5.0, partially purified enzymes have half-lives of several weeks to months
0C, suspension of crystals, 10 mM glycerol, 1 mM EDTA, 1 mM 2-mercaptoethanol, 0.1 M potassium phosphate, pH 7.0, saturated with ammonium sulfate, stable for several years
-
4C, crystallized suspension in 2.2 M ammonium sulfate, stable for several months
Candida mycoderma
-
as crystalline suspension in saturated ammonium sulfate, solutions containing 10 mM glycerol, 1 mM EDTA and a thiol e.g. 2-mercaptoethanol, yeast enzyme stable for several months, E. coli enzyme for several years
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
application of triazine dye affinity chromatography to large-scale purification
-
GSTrap FF column chromatography, gel filtration
-
HiTrapQ HP column, Bio-Scale CHT20-I column, all purification procedures are carried out at 277 K.
-
metal-chelate affinity chromatography using a Ni-NTA column, anion exchange chromatography using a Mono Q 5/50 GL column and gel filtration chromatography using s Superdex 200 10/30 GL column,all purification steps are performed in standard buffer (20 mM TrisHCl (pH 7.5), 10 mM glycerol, 1 mM beta-mercaptoethanol) at 4C excluding the affinity chromatography purification; mutant enzyme G230D
-
native and mutant enzyme
-
Ni-NTA agarose column chromatography and Superdex 200 gel filtration
nickel affinity column chromatography
normal and genetically altered enzyme
-
partial
recombinant enzyme
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
both isoforms expressed in COS-7 cells
contains PTS1-like targeting sequence for glycosomal localization
-
expressed in Escherichia coli
expressed in Escherichia coli as a fusion protein with maltose-binding protein
expressed in Escherichia coli BL21(DE3)pLysS cells
-
expressed in Escherichia coli JM109 (DE3 + pRARE2) cells
expressed in Escherichia coli Rosetta2 (DE3) pLysS cells
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expressed in Escherichia coli Shot BL21 star (DE3) cells
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expressed in H4IIE cell rat hepatoma cells
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expressed in Pichia pastoris X-33 cells
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His-tag, expressed in Escherichia coli
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overproduced in Escherichia coli BL21 (DE3)
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
in ambient smelt, activity remains constant until April when it starts to increase significantly to reach a maximum in May at levels almost 25times higher than those recorded in November
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transcript is upregulated by addition of glycerol
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upregulated in diapause eggs exposed to 5C
upregulated in sexual blood stage parasite
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
S414N
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increased thermostability, mechanism of stabilization
H232E
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residue located in the activation loop
H232R
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residue located in the activation loop, mutant protein has enhanced activity
D72V <