Information on EC 1.1.1.9 - D-xylulose reductase

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

EC NUMBER
COMMENTARY hide
1.1.1.9
-
RECOMMENDED NAME
GeneOntology No.
D-xylulose reductase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
xylitol + NAD+ = D-xylulose + NADH + H+
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
redox reaction
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
degradation of sugar alcohols
-
-
Metabolic pathways
-
-
Pentose and glucuronate interconversions
-
-
xylitol degradation
-
-
SYSTEMATIC NAME
IUBMB Comments
xylitol:NAD+ 2-oxidoreductase (D-xylulose-forming)
Also acts as an L-erythrulose reductase.
CAS REGISTRY NUMBER
COMMENTARY hide
9028-16-4
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain Y25
-
-
Manually annotated by BRENDA team
strain Y25
-
-
Manually annotated by BRENDA team
gene xdhA
-
-
Manually annotated by BRENDA team
strain F-3
-
-
Manually annotated by BRENDA team
strain F-3
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
CBS 4435
-
-
Manually annotated by BRENDA team
Candida tenuis CBS 4435
CBS 4435
-
-
Manually annotated by BRENDA team
strain Y-456
-
-
Manually annotated by BRENDA team
isolated from soil, China
-
-
Manually annotated by BRENDA team
strain Y-488
-
-
Manually annotated by BRENDA team
strain Y-488
-
-
Manually annotated by BRENDA team
FTI 20037
-
-
Manually annotated by BRENDA team
strain Y-1017
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
strain CFN42
UniProt
Manually annotated by BRENDA team
strain Y-1632
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
metabolism
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
D-arabitol + NAD+
?
show the reaction diagram
-
-
-
-
?
D-arabitol + NAD+
? + NADH + H+
show the reaction diagram
D-fructose + NADH + H+
D-sorbitol + NAD+
show the reaction diagram
-
-
-
-
-
D-iditol + NAD+
?
show the reaction diagram
D-mannitol + NAD+
? + NADH + H+
show the reaction diagram
8% of the activity with xylitol
-
-
?
D-mannitol + NAD+
D-fructose + NADH + H+
show the reaction diagram
D-ribitol + NAD+
D-ribulose + NADH
show the reaction diagram
D-ribulose + NADH
D-ribitol + NAD+
show the reaction diagram
-
-
-
r
D-ribulose + NADH + H+
?
show the reaction diagram
-
-
-
?
D-sorbitol + NAD+
D-fructose + NADH + H+
show the reaction diagram
D-sorbitol + NAD+
L-sorbose + NADH + H+
show the reaction diagram
D-threitol + NAD+
?
show the reaction diagram
D-xylitol + NAD+
D-xylulose + NADH
show the reaction diagram
D-xylulose + NAD+
xylitol + NADH + H+
show the reaction diagram
-
-
-
-
r
D-xylulose + NADH + H+
D-xylitol + NAD+
show the reaction diagram
D-xylulose + NADH + H+
xylitol + NAD+
show the reaction diagram
dihydroxyacetone + NADH
glycerol + NAD+
show the reaction diagram
-
-
-
-
-
erythritol + NAD+
? + NADH + H+
show the reaction diagram
0.77% of the activity with xylitol
-
-
?
erythritol + NAD+
L-erythrulose + NADH
show the reaction diagram
-
-
-
r
galactitol + NAD+
? + NADH + H+
show the reaction diagram
0.98% of the activity with xylitol
-
-
?
glycerol + NAD+
?
show the reaction diagram
L-arabinitol + NAD+
? + NADH + H+
show the reaction diagram
21% of the activity with xylitol
-
-
?
L-arabitol + NAD+
?
show the reaction diagram
L-erythrulose + NADH
erythritol + NAD+
show the reaction diagram
L-iditol + NAD+
?
show the reaction diagram
-
-
-
-
?
L-sorbose + NADH + H+
?
show the reaction diagram
-
-
-
?
L-threitol + NAD+
? + NADH + H+
show the reaction diagram
L-xylulose + NADH
L-xylitol + NAD+
show the reaction diagram
-
-
-
-
?
meso-erythritol + NAD+
? + NADH + H+
show the reaction diagram
ribitol + NAD+
?
show the reaction diagram
-
-
-
?
ribitol + NAD+
? + NADH + H+
show the reaction diagram
40% of the activity with xylitol
-
-
?
sorbitol + NAD+
D-fructose + NADH + H+
show the reaction diagram
-
-
-
-
?
Xylitol + NAD+
?
show the reaction diagram
xylitol + NAD+
D-xylose + NADH + H+
show the reaction diagram
xylitol + NAD+
D-xylulose + NADH + H+
show the reaction diagram
xylitol + NAD+
L-xylulose + NADH + H+
show the reaction diagram
xylitol + NADP+
D-xylulose + NADPH + H+
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
Xylitol + NAD+
?
show the reaction diagram
xylitol + NAD+
D-xylulose + NADH + H+
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ca2+
1 mM, 18% of the activity with Mg2+
Co2+
1 mM, 38% of the activity with Mg2+
Cu2+
1 mM, 51% of the activity with Mg2+
KCl
-
slight activation
MnCl2
-
10 mM, increase of activity by 100%
NaCl
-
activation
NH4Cl
-
activation
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
cysteine
-
at 294 mM
D-xylulose
HgCl2
-
complete inhibition
iodoacetate
-
complete inhibition
MgCl2
-
10 mM, 4% inhibition
NaCl
-
at 160 mM
NH4Cl
-
at 310 mM
NiCl2
-
1 mM, complete inhibition
Pb(CH3COO)2
-
-
PCMB
-
complete inhibition
additional information
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
cysteine
D-glucose
-
induces activity
D-Lyxose
-
induces activity
D-xylose
glycerol
-
induces activity
glycine
-
slight activation
GSH
-
activation
L-arabinose
-
induces activity
Sodium acetate
-
1 g/l in reaction medium, about 15% increase of activity
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.7
D-ribulose
35C, pH 7.5, native enzyme
4 - 30
D-sorbitol
12 - 94
D-xylitol
0.66 - 10
D-xylulose
0.9
L-sorbose
35C, pH 7.5, native enzyme
0.038 - 500
NAD+
0.037 - 0.24
NADH
0.0205 - 170
NADP+
50 - 496
ribitol
0.16 - 785
sorbitol
0.039 - 111
xylitol
13.8
xylulose
-
-
additional information
additional information
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
920
D-fructose
Candida sp. HA 167
-
25C, pH 7.5, Tris-HCl
74
D-ribulose
Blastobotrys adeninivorans
Q6KAV2
35C, pH 7.5, native enzyme
114 - 528
D-sorbitol
302.9 - 1800
D-xylulose
216
L-sorbose
Blastobotrys adeninivorans
Q6KAV2
35C, pH 7.5, native enzyme
0.052 - 30.33
NAD+
0.017 - 210
NADP+
1.19 - 161
sorbitol
17.9 - 2644
xylitol
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
10
D-xylulose
-
-
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.005
ZnSO4
Candida sp. HA 167
-
value identical for wildtype and for mutant E154C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.15 - 0.18
-
depending on nitrogen source in culture medium
0.76
-
purified native enzyme, pH 11.0, 30C, substrate meso-erythritol
2.16
-
specific activity of recombinant Saccharomyces cerevisiae strain TMB3057
6.74
-
purified native enzyme, pH 11.0, 30C, substrate D-arabitol
12.89
-
purified native enzyme, pH 11.0, 30C, substrate D-mannitol
102.5
-
purified native enzyme, pH 11.0, 30C, substrate xylitol
113
-
substrate D-xylitol, pH 10, 30C
156.1
-
purified native enzyme, pH 11.0, 30C, substrate D-sorbitol
180.4
-
purified native enzyme, pH 5.0, 30C, substrate D-fructose
220
-
substrate D-sorbitol, pH 10, 30C
250
-
pH 8.4, 30C
564
-
purified native enzyme, pH 5.0, 30C, substrate D-xylulose
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5
-
reduction of D-xylulose
5.5
-
reduction reaction
6.7
-
reduction of erythrulose
7.2
-
xylitol formation
7.5
native and recombinant enzyme
9.1 - 10
-
substrate oxidation, plateau
10.5 - 11
-
oxidation reaction
11
-
oxidation of xylitol
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
8.7 - 10.5
-
about 80% of maximal oxidation activity at pH 8.7 and 10.5
9 - 10
-
fully active
9 - 12
-
oxidation reaction, activity range
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20 - 50
-
20C: about 50% of maximal activity, 50C: about 50% of maximal activity
20 - 55
-
activity range, profile overview
35 - 68
-
about half-maximal activity at 35C and 68C
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6
-
isoelectric focusing
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
additional information
-
the organism grows on rice straw hemicellulosic hydrolysate, as the only source of nutrient, optimization of culture conditions for production of xylitol from D-xylose, xylitol dehydrogenase remains constant, whereas the level of xylose reductase decreases when the initial xylose concentration is increased from 30 to 70 g/l, development of enzyme activities, overview
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
additional information
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
72000
native enzyme, gel filtration, non-denaturing PAGE
80000
recombinant enzyme, gel filtration, non-denaturing PAGE
82000
-
gel filtration
94000
-
PAGE
120000
-
gel filtration
130000
-
PAGE
135000
gel filtration
142000
-
PAGE
160000
172000
-
gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homotetramer
tetramer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
hanging drop vapor diffusion method, crystal structure of the holoenzyme to 1.9 A resolution
multi-template homology modeling. The structural model of XDH obtained consists of a classical alpha/beta Rossmann fold pattern commonly found in the MDR family, which is organized into two beta-barrel domains, the coenzyme-binding residues 163-300 and catalytic residues 1-162, 301-364
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 7
-
24 h, fully stable
655092
6.8 - 7.4
-
90% of maximal activity retained at pH 6.8 and 7.4 at 2C
246415
7
-
24 h stable at 2C
246415
7 - 9
-
stable
246421
7.2 - 7.9
stable
667227
8 - 10
-
4C, 60 min, stable
686433
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0 - 40
-
10 min, 2.5 mM NAD+, stable
20
-
at room temperature, the enzyme is inactivated within 2 days
25
-
and below stable
30
-
1 h, completely stable
40
half-life 120 min
53.1
half denaturation temperature
additional information
-
wild-type: half denaturation temperature T1/2 (C): 35.2, thermal transition temperature Tcd (C): 43.0
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
2-mercaptoethanol stabilizes during purification
-
30C, pH 6-7, 24 h, fully stable
-
crude water extract from liver acetone-powder unstable
-
EDTA stabilizes during purification
-
freezing/thawing inactivates, 8 cycles lead to 89% loss of activity and 11 cycles to complete inactivation
ORGANIC SOLVENT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Acetone
-
stable to precipitation with 50% v/v
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-10C, acetone precipitate stable for weeks
-
-15C, 60% loss of activity after 1 month
-
-20C, stable with 50% v/v glycerol
-
-2C, purified stable for a month
-
0C, 34% loss of activity within a month
-
0C, partially purified stable for weeks
-
2C, 90% of maximal activity retained at pH 6.8 and 7.4 after 24 h
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
native enzyme 469fold from sstrain NH-10 by ammonium sulfate fractionation, two different steps of anion exchange chromatography, and gel filtration
-
optimized extraction by cetyl trimethyl ammonium bromide reversed micelles
-
partial
recombinant enzmye
recombinant enzyme
using affinity chromatography and preparative gel filtration
-
using Ni-NTA chromatography
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed as a His-tagged fusion protein in Escherichia coli
-
expressed in Escherichia coli
-
expressed in Escherichia coli JM 109
-
expressed in transketolase-deficient Saccharomyces cerevisiae strain (W303/tkl1 tkl2/2C)
-
expression in Escherichia coli
expression of the engineered D202A/L203R/V204S/E205P/S206R mutant enzyme fron Galactocandida mastotermitis with altered cofactor specificity, co-expression with a mutant NADPH-specific xylulose reductase from Candida tenuis in Saccharomyces cerevisiae, the transformed strain shows up to 50% decreased glycerol yield without increase in ethanol during xylose fermentation, overview
-
gene xdhA, coexpression of gene xdhA and cofactor regeneration enzyme, i.e. glucose dehydrogenase gene gdh from Bacillus subtilis, in Escherichia coli strain BL21(DE3), subcloning in Escherichia coli strain JM109
-
genes XYL2 (D207A/I208R/F209S) and XYL2 (S96C/S99C/Y102C/D207A/I208R/F209S) are introduced into Saccharomyces cerevisiae, which already contain the Pichia stipitis XYL1 gene (encoding xylose reductase) and the endogenously overexpressed XKS1 gene (encoding xylulokinase)
-
overexpression in Saccharomyces cerevisiae
-
overexpression of the enzyme in Saccharomyces cerevisiae strain CEN.PK 113-7D under control of the constitutive TDH3 promoter, co-expression with xylose reductase from Candida tenuis
-
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D202A/L203R/V204S/E205P/S206R
-
site-directed mutagenesis, introduction of multiple site-directed mutations in the coenzyme-binding pocket of Galactocandida mastotermitis XDH to enable activity with NADP+, which is lacking in the wild-type enzyme, genetic metabolic engineering for improvement of xylose metabolism and fermentation in wild-type Saccharomyces cerevisiae strains, which are not able to naturally metabolize D-xylulose, overview
E154C
-
mutant bearing a disrupted Zn2+ binding site: purified preparations show a variable Zn2+ (0.10-0.40 atom/subunit), mutant exhibits a constant catalytic Zn2+ centre activity and does not require exogenous Zn2+ for activity or stability. E154C retains 0.019% and 0.74% of wild-type catalytic efficiency (kcat/Km (sorbitol): 7800/Msec and kcat:161/sec) for NAD+-dependent oxidation of sorbitol at 25C respectively. The pH profile of kcat/Ksorbitol for E154C decreases below an apparent pK of 9.1, reflecting a shift in pK by about +1.7-1.9 pH units compared with the corresponding pH profiles for wild-type. IC50 (ZnSO4): 0.005 mM
synthesis
-
use of enzyme in a process for producing xylitol from D-glucose
D38S/M39R
the mutant enzyme is able to exclusively use NADP+, with no loss of activity
D207A
kcat/Km for NAD+ is 3.6fold lower than wild-type value, kcat/Km for NADP+ is 4.3fold higher than wild-type value
D207A/F209S
kcat/Km for NAD+ is 2.2fold lower than wild-type value, kcat/Km for NADP+ is 745fold higher than wild-type value
D207A/I208R
kcat/Km for NAD+ is 2.5fold lower than wild-type value, kcat/Km for NADP+ is 229fold higher than wild-type value
D207A/I208R/F209S
D207A/I208R/F209S/N211R
kcat/Km for NAD+ is 32.9fold lower than wild-type value, kcat/Km for NADP+ is 4292fold higher than wild-type value, increased thermostability
D207A/I208R/F209T
kcat/Km for NAD+ is 2.4fold lower than wild-type value, kcat/Km for NADP+ is 4754fold higher than wild-type value
D207A/I208R/F209Y
kcat/Km for NAD+ is 6.9fold lowerthan wild-type value, kcat/Km for NADP+ is 788fold higher than wild-type value
F209S
kcat/Km for NAD+ is 1.9fold lower than wild-type value, kcat/Km for NADP+ is 31.4fold higher than wild-type value
I208R
kcat/Km for NAD+ is nearly identical to wild-type value, kcat/Km for NADP+ is 44fold higher than wild-type value
I208R/F209S
kcat/Km for NAD+ is 30.7fold lower than wild-type value, kcat/Km for NADP+ is 1.5fold higher than wild-type value
N211R
kcat/Km for NAD+ is 1.1fold lower than wild-type value, kcat/Km for NADP+ is 7.6fold higher than wild-type value
S96C/S99C/Y102C
-
specific activity (U/min): 1440, half denaturation temperature T1/2 (C): 46.1, thermal transition temperature Tcd (C): 47.5
S96C/S99C/Y102C/D207A/I208R/F209S
S96C/S99C/Y102C/D207A/I208R/F209S/N211R
kcat/Km for NAD+ is 26.5fold lower than wild-type value, kcat/Km for NADP+ is 16154fold higher than wild-type value
S96C/S99C/Y102C/E101F
-
specific activity (U/min): 1550, half denaturation temperature T1/2 (C): 50.9, thermal transition temperature Tcd (C): 50.5
S96C/S99C/Y102C/F98R
-
specific activity (U/min): 1510, half denaturation temperature T1/2 (C): 53.1, thermal transition temperature Tcd (C): 51.7
S96C/S99C/Y102C/F98R/E101F
-
specific activity (U/min): 1620, half denaturation temperature T1/2 (C): 56.0, thermal transition temperature Tcd (C): 53.8
S96C/S99C/Y102C/H112D
-
specific activity (U/min): 1360, half denaturation temperature T1/2 (C): 44.0, thermal transition temperature Tcd (C): 47.0
S96C/S99C/Y102C/P95S
-
specific activity (U/min): 1220, half denaturation temperature T1/2 (C): 37.4, thermal transition temperature Tcd (C): 43.5
S96C/S99CY102C
kcat/Km for NAD+ is 1.1fold lower than wild-type value, kcat/Km for NADP+ is 8.8fold higher than wild-type value
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
biotechnology
molecular biology
-
a high thermostability of PsXDH is obtained by subsequent site-directed mutagenesis of the structural zinc-binding loop. The best mutant in this study (C4/F98R/E101F) shows a 10.8 C higher thermal transition temperature and 20.8 C higher half denaturation temperature (T1/2) compared with wild-type
synthesis
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