Information on EC 5.3.1.5 - xylose isomerase and Organism(s) Streptomyces rubiginosus and UniProt Accession P24300

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Streptomyces rubiginosus
UNIPROT: P24300


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


The taxonomic range for the selected organisms is: Streptomyces rubiginosus

EC NUMBER
COMMENTARY hide
5.3.1.5
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RECOMMENDED NAME
GeneOntology No.
xylose isomerase
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
isomerization
-
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
xylose degradation I
-
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d-xylose degradation
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Pentose and glucuronate interconversions
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Fructose and mannose metabolism
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Metabolic pathways
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SYSTEMATIC NAME
IUBMB Comments
D-xylose aldose-ketose-isomerase
Contains two divalent metal ions, preferably magnesium, located at different metal-binding sites within the active site. The enzyme catalyses the interconversion of aldose and ketose sugars with broad substrate specificity. The enzyme binds the closed form of its sugar substrate (in the case of glucose, only the alpha anomer) and catalyses ring opening to generate a form of open-chain conformation that is coordinated to one of the metal sites. Isomerization proceeds via a hydride-shift mechanism.
CAS REGISTRY NUMBER
COMMENTARY hide
9023-82-9
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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-glucose
alpha-D-fructose
show the reaction diagram
-
-
-
-
r
D-glyceraldehyde
dihydroxyacetone
show the reaction diagram
-
-
-
-
?
D-Xylose
D-Xylulose
show the reaction diagram
-
-
-
-
r
D-Glucose
D-Fructose
show the reaction diagram
D-Xylose
D-Xylulose
show the reaction diagram
L-Arabinose
L-Ribulose
show the reaction diagram
low reaction efficiency
-
-
r
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
D-Xylose
D-Xylulose
show the reaction diagram
-
-
-
-
r
D-Xylose
D-Xylulose
show the reaction diagram
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Co2+
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activates
Fe2+
-
activates
Mn2+
-
activates
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
D-glyceraldehyde
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competitive inhibition of the isomerization of D-xylose
D-Threonohydroxamic acid
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competitive inhibitor, complete inhibition at 0.5 mM
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
3
D-glyceraldehyde
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in D2O, 24 mM imidazole buffer, 10 mM MgCl2, at pH 7.5 and 25°C
4.9
D-xylose
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in D2O, 24 mM imidazole buffer, 10 mM MgCl2, at pH 7.5 and 25°C
160 - 340
D-glucose
1 - 548
D-xylose
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0001
D-glyceraldehyde
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in D2O, 24 mM imidazole buffer, 10 mM MgCl2, at pH 7.5 and 25°C
2.4
D-xylose
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in D2O, 24 mM imidazole buffer, 10 mM MgCl2, at pH 7.5 and 25°C
0.03 - 9.22
D-glucose
0.007 - 5.52
D-xylose
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.000034
D-glyceraldehyde
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in D2O, 24 mM imidazole buffer, 10 mM MgCl2, at pH 7.5 and 25°C
0.49
D-xylose
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in D2O, 24 mM imidazole buffer, 10 mM MgCl2, at pH 7.5 and 25°C
0.00013 - 1.84
D-xylose
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
8 - 8.2
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in presence of Mg2+ and Co2+, mutants D65A and D163N/E167Q
8
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in presence of Co2+, mutant E221A; in presence of Mg2+ and Co2+, mutants E221A and D56N
8.1 - 8.3
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in presence of Mg2+ and Co2+, mutant D81A
8.5
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in presence of Mg2+ and Co2+, wild-type
8.8
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in presence of Co2+, wild-type
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
PDB
SCOP
CATH
UNIPROT
ORGANISM
Streptomyces rubiginosus;
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
160000
-
neutron diffraction
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homotetramer
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x-ray crystallography
tetramer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
in complex with D-glyceraldehyde, hanging drop vapor diffusion method, using 0.2-0.3 M Mg-formate at pH 7.0 and 22°C
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analysis of the location of hydrogen atoms by time-of-flight neutron Laue technique. The neutron structure of crystalline XI with bound product, D-xylulose, shows, that O5 of D-xylulose is not protonated but is hydrogen-bonded to doubly protonated His54. Also, Lys289, which is neutral in native XI, is protonated, while the catalytic water in native XI has become activated to a hydroxyl anion which is in the proximity of C1 and C2, the molecular site of isomerization of xylose
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cooling crystallization from 0.17 M MgSO4 solution
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neutron diffraction, largest crystals at 18°C, 95 mg/ml xylose isomerase, 16.9% ammonium sulfate, mathematical analysis to determine optimal conditions for crystallization
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neutron quasi-Laue diffraction, resolution: 2.2 A - clear visibility of deuterium atoms, clarification of critical residues at the active site and their protonation states
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study on the dynamics of solute transport in orthorhombic D-xylose isomerase crystals by means of Brownian dynamics and molecular dynamics simulations and investigation of the diffusion of S-phenylglycine molecules inside XI crystals. The S-phenylglycine molecules mostly interact with residues His54, Asp287, and Lys183. In general, the diffusivities of solute species are found to be 1 to 2 orders of magnitude lower than those of the corresponding free molecules in water
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time-of-flight neutron diffraction at 1.8 A resolution, metal-free enzyme - emphasis on the active site of xylose isomerase, especially of protonation states of His, Lys and H2O
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ORGANIC SOLVENT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Acetone
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70-80% of cross-linked crystalline xylose isomerase activity is left after incubation for 24 h at 50°C in buffer solutions (pH 7.2) containing 10-90% acetone. Soluble xylose isomerase is considerably less stable in acetone-containing solutions. The addition of acetone enhances the production of fructose from glucose by enhancing the reaction rate and shifting the equilibrium toward fructose. However, xylose isomerase must be in the form of cross-linked crystals for maximal activity and stability
Ethanol
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in buffer containing 50% ethanol only 2% of the initial cross-linked crystalline xylose isomerase activity is retained after 24 h at 50°C. Soluble xylose isomerase is considerably less stable in ethanol-containing solutions
Methanol
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cross-linked crystalline xylose isomerase is inactivated in 24 h at 50°C even more in maleate buffer containing 50% methanol
Pyridine
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cross-linked crystalline xylose isomerase is inactivated in 24 h at 50°C even more in maleate buffer containing 10% pyridine
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
from a stock solution of a food grade product, 2 counterdiffusion dialysis devices: diffusion-controlled apparatus for microgravity and counterdiffusion cell, growth of crystals approx. 3 months
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gel filtration over Sephacryl S200 - dialysis against 0.05 g/l MgSO4, purity control verified with capillary electrophoresis
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gel filtration with Sephacryl S-200 high resolution media, running buffer composed of 0.05 M sodium phosphate pH 7.7, 0.1 M sodium chloride and 0.02% azide - final purity of xylose isomerase: 96%
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HisTrap crude nickel affinity column chromatography
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli BL21(DE3) cells
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expression in Escherichia coli
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D163N/E167Q
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40-60% of wild-type activity, lower pH optimum than wild-type, nearly same tehrmostability as wild-type
D287N
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inactive
D56N
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turnover number increased by 30-40% over that ofwild-type at pH 7.3, lower pH optimum than wild-type, nearly same thermostability as wild-type
D65A
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40-60% of wild-type activity, lower pH optimum than wild-type, nearly same tehrmostability as wild-type
D81A
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40-60% of wild-type activity, lower pH optimum than wild-type, nearly same tehrmostability as wild-type
E221A
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turnover number increased by 30-40% over that from wild-type at pH 7.3, lower pH optimum than wild-type, nearly same tehrmostability as wild-type
H220S
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decreased affinity for Mg2+ and decraesed activity in contrast to wild-type
N185K
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decreased affinity for Mg2+ and decraesed activity in contrast to wild-type
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
nutrition
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high fructose corn syrups
synthesis