Information on EC 3.5.99.6 - glucosamine-6-phosphate deaminase

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

EC NUMBER
COMMENTARY hide
3.5.99.6
-
RECOMMENDED NAME
GeneOntology No.
glucosamine-6-phosphate deaminase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
alpha-D-glucosamine 6-phosphate + H2O = D-fructose 6-phosphate + NH3
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
group transfer
-
-
transmolecular
-
intramolecular oxidoreduction
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Amino sugar and nucleotide sugar metabolism
-
-
chitin degradation I (archaea)
-
-
chitin derivatives degradation
-
-
Metabolic pathways
-
-
N-acetylglucosamine degradation I
-
-
UDP-N-acetyl-D-galactosamine biosynthesis II
-
-
metabolism of amino sugars and derivatives
-
-
SYSTEMATIC NAME
IUBMB Comments
2-amino-2-deoxy-D-glucose-6-phosphate aminohydrolase (ketol isomerizing)
The enzyme uses ring opening and isomerization of the aldose-ketose type to convert the -CH(-NH2)-CH=O group of glucosamine 6-phosphate into -C(=NH)-CH2-OH, forming 2-deoxy-2-imino-D-arabino-hexitol, which then hydrolyses to yield fructose 6-phosphate and ammonia. N-Acetyl-D-glucosamine 6-phosphate, which is not broken down, activates the enzyme.
CAS REGISTRY NUMBER
COMMENTARY hide
9013-10-9
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain A158
-
-
Manually annotated by BRENDA team
strain A158
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
gene glmD
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
strain UA159, gene smu.636
-
-
Manually annotated by BRENDA team
strain UA159, gene smu.636
-
-
Manually annotated by BRENDA team
hog
-
-
Manually annotated by BRENDA team
strain KOD1
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
-
mutant parasites lacking GND are unable to grow in medium containing amino sugars as sole carbohydrate source and rapidly loose viability, concomitant with the hyper-accumulation of hexosamine-phosphates. Expression of native GND, but not a cytosolic form of GND, in DELTAgnd parasites restored hexosamine-dependent growth, indicating that toxicity is due to depletion of glycosomal pools of ATP. Promastigote and amastigote stages of the DELTAgnd mutant are unable to replicate within macrophages and were either completely cleared or exhibited reduced lesion development in highly susceptible Balb/c mice
physiological function
hGNPDA1 can be important for the maintenance of an adequate level of the pool of the UDP-GlcNAc6P, the N-acetylglucosylaminyl donor for many reactions in the cell
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
alpha-D-glucosamine 6-phosphate + H2O
D-fructose 6-phosphate + NH3
show the reaction diagram
D-fructose 6-phosphate + NH3
alpha-D-glucosamine 6-phosphate + H2O
show the reaction diagram
-
-
-
-
r
D-fructose 6-phosphate + NH3
D-glucosamine 6-phosphate + H2O
show the reaction diagram
-
deamination reaction is preferred over amination/isomerization
-
-
r
D-glucosamine 6-phosphate + H2O
D-fructose 6-phosphate + NH3
show the reaction diagram
glucosamine 6-phosphate + H2O
fructose 6-phosphate + NH3
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
alpha-D-glucosamine 6-phosphate + H2O
D-fructose 6-phosphate + NH3
show the reaction diagram
D-fructose 6-phosphate + NH3
alpha-D-glucosamine 6-phosphate + H2O
show the reaction diagram
-
-
-
-
r
D-glucosamine 6-phosphate + H2O
D-fructose 6-phosphate + NH3
show the reaction diagram
glucosamine 6-phosphate + H2O
fructose 6-phosphate + NH3
show the reaction diagram
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
arsenate
-
activates
Co2+
-
activates
Hg2+
-
activates
Mg2+
-
activates
Mn2+
-
activates, reverses inhibition by EDTA
Ni2+
-
activates
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2,5-anhydro mannitol 6-phosphate
-
-
2-deoxy-2-amino-D-glucitol 6-phosphate
5,5'-dithiobis(2-nitrobenzoate)
-
-
Cd2+
-
-
Cu2+
-
-
cysteine
-
-
D-fructose 1,6-diphosphate
-
-
D-fructose 6-phosphate
D-fructose oxime 6-phosphate
-
-
D-Glucosamine 6-phosphate
-
substrate inhibition
D-glucose 6-phosphate
-
-
diethyl dicarbonate
-
complete inactivation, 2-deoxy-2-amino-D-glucitol 6-phosphate protects
diphosphate
-
-
Fluorescamine
-
-
N-acetylglucosamine 6-phosphate
-
-
N3-(4-Methoxyfumaroyl)-L-2,3-diaminopropanoic acid
-
-
Ni2+
-
-
phosphate
-
slight inhibition
UDP-N-acetylglucosamine
-
slight inhibition at 1.67 mM, slight activation at 0.00167-0.167 mM
additional information
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2-deoxyglucose 6-phosphate
-
activates
6-phosphogluconate
-
activates
D-glucose 6-phosphate
D-mannose 6-phosphate
-
-
N-acetylgalactosamine
-
-
N-Acetylgalactosamine 6-phosphate
-
activates
N-acetylglucosamine
-
-
N-acetylglucosamine 6-phosphate
N-acetylglucosamine-6-phosphate
-
allosteric activator of NagB
PCMB
-
inhibits at high concentrations, activates at low concentrations
UDP-N-acetylglucosamine
-
slight activation at 0.00167-0.167 mM, slight inhibition at 1.67 mM
additional information
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.21 - 45.2
alpha-D-glucosamine 6-phosphate
1.1 - 36
D-fructose 6-phosphate
0.041 - 17
D-Glucosamine 6-phosphate
1.7
fructose 6-phosphate
-
amination reaction, pH 8.0, 30C
5
glucosamine 6-phosphate
-
deamination reaction, pH 8.0, 30C
2 - 41.6
NH3
3.7 - 300
NH4+
additional information
additional information
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
107 - 158
alpha-D-glucosamine 6-phosphate
5.8 - 455
D-fructose 6-phosphate
0.044 - 1800
D-Glucosamine 6-phosphate
39.6
NH3
Thermococcus kodakarensis
-
pH 7.3, 60C
additional information
additional information
-
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
13.4
2,5-anhydro mannitol 6-phosphate
-
pH 7.7, 30C
0.0041
2-deoxy-2-amino D-glucitol 6-phosphate
-
pH 7.7, 30C
0.002 - 0.0043
2-deoxy-2-amino-D-glucitol 6-phosphate
1.3
D-fructose oxime 6-phosphate
-
pH 7.7, 30C
4
D-Glucosamine 6-phosphate
-
22C, pH 8.0
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.036
-
initial culture, cell homogenate
0.5
-
after 7.2fold purification, HisC-tagged enzyme, with D-fructose 6-phosphate as substrate, at pH 7.0 and 37C
8.6
-
crude extract, with D-fructose 6-phosphate as substrate, at pH 7.0 and 37C
62
-
after 7.2fold purification, untagged wild type enzyme, with D-fructose 6-phosphate as substrate, at pH 7.0 and 37C
73
-
after 7.2fold purification, HisN-tagged enzyme, with D-fructose 6-phosphate as substrate, at pH 7.0 and 37C
85
-
purified enzyme
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.5 - 8
-
-
6.6
-
in absence of activator
7 - 8.2
-
-
7.1 - 7.3
-
in absence of activators
7.4
-
in presence of D-glucose 6-phosphate
7.5
-
enzyme activated by N-acetylgalactosamine 6-phosphate
8 - 8.5
-
both amination and deamination reaction
8.2 - 8.4
-
in presence of phosphate and D-glucose 6-phosphate
8.5 - 8.7
-
-
8.9
-
both directions
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 10
-
pH-profile, overview
6 - 8.5
-
pH 6.0: about 25% of maximal activity, pH 8.5: about 60% of maximal activity
6.2 - 8
-
pH 6.2: about 25% of maximal activity, pH 8.0: about 55% of maximal activity, in presence of glucose 6-phosphate
6.8 - 8.3
-
70% of maximal activity at pH 6.8, maximal activity at pH 8.3
7 - 9.5
-
pH 7.0: about 55% of maximal activity, pH 9.5: about 74% of maximal activity
additional information
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
32 - 47
-
about 90% of maximal activity at 32C and 47C
additional information
-
effect of temperature on homotropic co-operativity in the enzyme
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.7
-
isoelectric focusing and chromatofocusing
5.9
-
electrofocusing on polyacrylamide gels
8.55
-
chromatofocusing
PDB
SCOP
CATH
ORGANISM
UNIPROT
Bacillus subtilis (strain 168)
Bacillus subtilis (strain 168)
Borrelia burgdorferi (strain ATCC 35210 / B31 / CIP 102532 / DSM 4680)
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)
Streptococcus mutans serotype c (strain ATCC 700610 / UA159)
Streptococcus mutans serotype c (strain ATCC 700610 / UA159)
Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
Vibrio cholerae serotype O1 (strain ATCC 39315 / El Tor Inaba N16961)
Vibrio cholerae serotype O1 (strain ATCC 39315 / El Tor Inaba N16961)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
17500
-
gel filtration
25500
-
x * 25500, recombinant His-tagged enzyme, SDS-PAGE
29000
-
gel filtration
30400
-
6 * 30400, SDS-PAGE
30700
-
x * 30700, native enzyme, SDS-PAGE
32500
-
1 * 32500, SDS-PAGE
33000
-
x * 33000, SDS-PAGE
37300
-
2 * 37300, SDS-PAGE
71800
-
gel filtration
154000
-
sucrose density gradient centrifugation
180000
-
about, gel filtration
185000
-
gel filtration
190000
-
pore-gradient gel electrophoresis, density gradient centrifugation
210000
-
gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
-
2 * 37300, SDS-PAGE
hexamer
monomer
monomer or homodimer or homotetramer
-
the recombinant and wild type enzyme exhibit heterogeneity of the quaternary structure and in solution exist in three interconvertible forms, namely, monomeric, homodimeric, and homotetrameric. The monomer/dimer/tetramer ratios depended on protein concentration and fall within the range from 72:27:1 to 39:23:38. The enzyme is fully active in each of the oligomeric structures
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
side-chain modification
-
enzyme is chemically transaminated, modifying its N-terminal methionine residue to a 2-oxo-4-(methylthio)butyryl group, the transamination markedly reduces the affinity of the enzyme for its allosteric activator N-acetylglucosamine 6-phosphate, in contrast with the unmodified enzyme, which behaves as a typical allosteric K-enzyme, the modified enzyme becomes a mixed K-V allosteric protein, borohydride reduction to obtain the corresponding enzyme with a terminal hydroxy group, this enzyme shows significant recovery of the catalytic catalytic activity and its allosteric activation pattern, becomes similar to that found for the unmodified enzyme; the modification by diethyl dicarbonate results in complete inactivation of enzyme
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
structure of the enzyme in complexes with its allosteric activator and with a competitive inhibitor
-
the crystallographic structure of F174A is determinated
-
the structure of five complexes of the enzyme in R and T state is described and their kinetic and allosteric implication analysed, the ligand-free enzyme, T-conformer, undergoes an allosteric transition to the free active-site R conformer upon binding of the allosteric activator
-
isozyme GNPDA1, X-ray diffraction structure determination and analysis at 1.75 A resolution
purified recombinant C-terminally His-tagged, selenomethionine-labeled enzyme, method optimization, sitting drop vapor diffusion, 0.002 ml of protein solution is mixed with 0.002 ml of reservoir solution containing 24% PEG 1000, 0.2 M sodium chloride, 0.1 M sodium/potassium phosphate, pH 6.25, cyroprotection by 10% glycerol, X-ray diffraction structure determination and analysis at 1.8-2.3 A resolution
-
purified recombinant His-tagged enzyme, the protein solution contains 10 mg/ml protein in 20 mM Tris-HCl, 150 mM NaCl, pH 7.5, mixing of 0.001 ml of protein and 0.001 ml of reservoir solution, and equilibration against 0.5 ml of reservoir solution, hanging drop vapour diffusion method, X-ray diffraction structure determination and anaylsis at 2.4 A resolution
-
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 9
-
4C, purified enzyme, overnight, in presence of different salts, stable
671576
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
60
-
10 min, in presence of 1.7 mM glucosamine 6-phosphate or N-acetylglucosamine 6-phosphate, 7% loss of activity; 90% loss of activity within 1 min without stabilizer, stable for 1 min in presence of 0.6 mM glucosamine 6-phosphate or N-acetylglucosamine 6-phosphate
65
-
complete loss of activity
80
-
half-life 103 min
90
-
half-life 19 min
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
D-glucosamine 6-phosphate and N-acetylglucosamine 6-phosphate stabilize against heat inactivation at 60C, and protect the enzyme from tryptic digestion
-
introduction of the oligoHis tag at the enzyme's C-terminus results in almost complete loss of the catalytic activity, while the catalytic properties of HisN-tagged enzyme and the untagged wild type are very similar
-
labile to freezing and thawing
protease-sensitive. Incubation with 0.3 mg/ml protease type V or 0.3 mg/ml trypsin inactivates completely
-
stable to dialysis
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-18C, little loss of activity after 2-3 months
-
-20C, 50% glycerol, stable for up to 2 months
-
-20C, in 150 mM Tris-HCl buffer, pH 7.25, 50% v/v glycerol, stable up to 6 months
-
-70C, stable for 70 days
-
0-5C, in liquid N2, little loss of activity after 2-5 months
-
0C, stable for at least 2 weeks
-
4C, purified enzyme, pH 5.0-9.0, overnight, in presence of different salts, stable
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate precipitation, Resource Q column chromatography, and Ni2+-IDA agarose column chromatography
-
N-6-aminohexanoyl-glucosamine 6-phosphate agarose column chromatography
-
native enzyme 1390fold from kidney cortex by affinity chromatography on N-epsilon-amino-n-hexanoyl-D-glucosamine-6-phosphate agarose and on phosphocellulose, the latter behaving as an active site affinity resin
-
native enzyme by ammonium sulfate fractionation, anion exchange chromatography, and affinity chromatography on a N-aminohexyl-glucosamine-6-phosphate resin folowed by dialysis and ultrafiltration
-
recombinant C-terminally His-tagged, selenomethionine-labeled enzyme from Escherichia coli strain B834 by heat treatment for 10 min at 85C, nickel affinity chromatography, and gel filtration
-
recombinant enzyme
recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) to homogeneity in two steps by nickel affinity chromatography and gel filtration
-
recombinant isozyme GNPDA1 from Escherichia coli strain IBPC 590 by single purification step by means of allosteric-site affinity chromatography on N-6-aminohexanoyl-GlcN6P agarose
wild-type enzyme and two mutants
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli BL21(DE3) pLysS cells
-
expressed in Escherichia coli strain LAA20
-
expressed in Saccharomyces cerevisiae strain BY
-
expression of isozyme GNPDA1 in Escherichia coli strain IBPC 590 (DELTAnagEBACD DELTAlac), which carries a deletion of the chromosomal copy of the nag operon and expresses the recombinant enzyme constitutively from the lac promoter due to the lacI mutation
expression of native GND, but not a cytosolic form of GND, in DELTAgnd parasites restores hexosamine-dependent growth. Targeted expression of glucosamine-6-phosphate deaminase, GND, in glycosomes via a canonical C-terminal targeting signal when expressed as a GFP fusion protein
-
gene glmD, overexpression of C-terminally His-tagged, selenomethionine-labeled enzyme in Escherichia coli strain B834
-
gene smu.636, cloning from genomic DNA, expression as His-tagged enzyme in Escherichia coli strain BL21(DE3)
-
two enzyme genes GNP1 and GNP2 in the completely sequenced genomes
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C118S/C228S/C239S
-
decrease in kcat-value, no modification of allosteric activation
C118Ser/C228S/C239S/D165C/S206W
-
the mutant shows reduced turnover number compared to the wild type enzyme
C219S
-
the kinetic and allosteric properties of the mutant enzyme in which Ser replaces Cys219 or Cys228 are the same as those described for the wild-type enzyme. The same result is obtained with the double mutation
C228S
-
the kinetic and allosteric properties of the mutant enzyme in which Ser replaces Cys219 or Cys228 are the same as those described for the wild-type enzyme. The same result is obtained with the double mutation
D141N
-
the mutation modifies the kcat versus pH profile of the enzyme
D141N/E148Q
-
mutation modifies the kcat versus pH profile of the enzyme
E148Q
-
the mutation modifies the kcat versus pH profile of the enzyme
H143Q
-
drastically impairs the activity of the enzyme in the forward but not in the backward direction of the reaction
K208E
-
entirely inactive in absence of allosteric activator N-acetylglucose-6-phosphate
K208V
-
homotropic cooperativity, Hill-coefficient 1.7, 2fold increase in dissociation constant for allosteric activator N-acetylglucose-6-phosphate compared to wild-type
R172A
-
inactive in absence of allosteric activator N-acetylglucose-6-phosphate, at high activator levels, cooperativity diminishes and substrate inhibition becomes significant
R172A/K208E
-
inactive in absence of allosteric activator N-acetylglucose-6-phosphate, at high activator levels, cooperativity diminishes and substrate inhibition becomes significant
W15Y
-
mutant containing a single Trp residue at W224
W15Y/F174W/W224Y
-
mutant containing a single, new Trp-residue at F174W
W15Y/W224Y
-
mutant without Trp residues
W15Y/W224Y/Y254W
-
mutant containing a single, new Trp-residue at Y254W
W224Y
-
mutant containing a single Trp residue at W15
Y121T
-
while the wild-type enzyme behaves as a classical allosteric K-system which can be described by the allosteric concerted model, the mutant forms Y121T and Y121W present an asymmetric behaviour towards the allosteric activator, which can be described as two distinct half-of-the-sites allosteric activation steps occuring with different affinities for the N-acetyl-D-glucosamine 6-phosphate
Y121W
-
while the wild-type enzyme behaves as a classical allosteric K-system which can be described by the allosteric concerted model, the mutant forms Y121T and Y121W present an asymmetric behaviour towards the allosteric activator, which can be described as two distinct half-of-the-sites allosteric activation steps occuring with different affinities for the N-acetyl-D-glucosamine 6-phosphate
Y254F
-
the mutant is less active than wild-type enzyme, the replacement causes an uncoupling of the homotropic and heterotrophic effects
Y254T
-
the mutation results in pure K-system with a similar catalytic activity to that of the wild-type enzyme, mutant displays kcat values similar to the wild-type enzyme
additional information
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