Information on EC 5.3.2.5 - 2,3-diketo-5-methylthiopentyl-1-phosphate enolase

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

EC NUMBER
COMMENTARY hide
5.3.2.5
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RECOMMENDED NAME
GeneOntology No.
2,3-diketo-5-methylthiopentyl-1-phosphate enolase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
5-(methylthio)-2,3-dioxopentyl phosphate = 2-hydroxy-5-(methylthio)-3-oxopent-1-enyl phosphate
show the reaction diagram
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Cysteine and methionine metabolism
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Metabolic pathways
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NIL
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S-methyl-5-thio-alpha-D-ribose 1-phosphate degradation
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SYSTEMATIC NAME
IUBMB Comments
2,3-diketo-5-methylthiopentyl-1-phosphate keto-enol-isomerase
The enzyme participates in the methionine salvage pathway in Bacillus subtilis [2]. In some species a single bifunctional enzyme, EC 3.1.3.77, acireductone synthase, catalyses both this reaction and EC 3.1.3.87, 2-hydroxy-3-keto-5-methylthiopentenyl-1-phosphate phosphatase [1].
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
planktonic organism
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Manually annotated by BRENDA team
no activity in Rhodospirillum rubrum
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Manually annotated by BRENDA team
a spontaneous streptomycin-resistant derivative of strain S1, gene rlpA
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
malfunction
metabolism
physiological function
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
2,3-diketo-1-phosphohexane
2-hydroxy-3-keto-l-phospho-l-hexene
show the reaction diagram
2,3-diketo-l-phospho-5-thiomethylpentane
?
show the reaction diagram
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-
-
-
?
5-(methylthio)-2,3-dioxopentyl 1-phosphate
2-hydroxy-3-oxo-5-methylthiopentenyl 1-phosphate
show the reaction diagram
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-
-
-
?
5-(methylthio)-2,3-dioxopentyl phosphate
2-hydroxy-5-(methylthio)-3-oxopent-1-enyl phosphate
show the reaction diagram
5-methylthio-2,3-dioxo-pentyl phosphate
2-hydroxy-5-methylthio-3-oxopentenyl phosphate
show the reaction diagram
5-methylthioribulose 1-phosphate
1-thiomethyl-D-xylulose 5-phosphate + 1-thiomethyl-D-ribulose 5-phosphate
show the reaction diagram
additional information
?
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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
2,3-diketo-1-phosphohexane
2-hydroxy-3-keto-l-phospho-l-hexene
show the reaction diagram
5-(methylthio)-2,3-dioxopentyl phosphate
2-hydroxy-5-(methylthio)-3-oxopent-1-enyl phosphate
show the reaction diagram
5-methylthio-2,3-dioxo-pentyl phosphate
2-hydroxy-5-methylthio-3-oxopentenyl phosphate
show the reaction diagram
5-methylthioribulose 1-phosphate
1-thiomethyl-D-xylulose 5-phosphate + 1-thiomethyl-D-ribulose 5-phosphate
show the reaction diagram
additional information
?
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
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no inhibition by phosphate
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.012 - 0.013
5-(methylthio)-2,3-dioxopentyl 1-phosphate
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
56 - 83
5-(methylthio)-2,3-dioxopentyl 1-phosphate
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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overall pathway activity, purified enzyme, pH 7.5, 30°C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25
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assay at
PDB
SCOP
CATH
ORGANISM
UNIPROT
Geobacillus kaustophilus (strain HTA426)
Geobacillus kaustophilus (strain HTA426)
Geobacillus kaustophilus (strain HTA426)
Geobacillus kaustophilus (strain HTA426)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25500
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gel filtration
28300
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1 * 28300, SDS-PAGE
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
additional information
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structure of the apo decarbamylated enzyme E form, computational structure analysis and modeling, structure comparison, overview. In the E form of DK-MTP-1P enolase the loop at 299-311, equivalent to loop-6 in RuBisCO, is in a closed conformation and the loop at 37-46, equivalent to the 60s loop, is positioned about 15 A away from the active site
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
activated wild-type enolase, carboxylated on Lys173, with Mg2+ alone, or Mg2+, and HCO3-, or Mg2+ and substrate 2,3-diketohexane 1-phosphate, and of the selenomethionine-substituted enzyme variant, hanging drop method, 10-15 mg/ml protein in 20 mM Tris-HCl, pH 7.9, 100 mM NaCl, and 5-10 mM MgCl2, 100 mM NaCl, and 1-5 mM NaHCO3, mixed with 24% PEG 3350, 0.1 M Tris-HCl, pH 8.5, and 0.2 M ammonium acetate as precipitant for the crystals of wild-type enzyme with Mg2+, or 25% PEG 3350, 0.1 M HEPES, pH 7.5, and 0.2 M ammonium acetate as precipitant for the other crystal variants, room temperature, 6 days, X-ray diffraction structure determination and analysis at 1.7 A resolution, molecular replacement
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
40
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the wild type enzyme shows no decline in activity after incubation at 40°C, and retains about 80% of its original activity after incubation at 45°C
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
His-Bind resin column chromatography and Superose 6 gel filtration
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native enzyme by fractionation with help of protamine suzlfate, ammonium sulfate, and P-60, followed by anion exchange and hydrophobic interaction chromatography
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recombinant enzyme from Escherichia coli
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recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography
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recombinant wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by two different steps of anion exchange chromatography and ultrafiltration
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Bacillus subtilis RLPalpha-1 sequence determination and analysis, and phylogenetic analysis
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expressed in Escherichia coli BL21 (DE3) pLysS cells
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expression of His-tagged enzyme in Escherichia coli strain BL21(DE3)
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expression of wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
gene rbcLIV, DNA and amino acid sequence determination and analysis, sequence comparisons and phylogenetic analysis, cloning in Escherichia coli strain JM109, expression in Escherichia coli strain BL21
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gene rlpA, form II RubisCO-like protein, cloning and expression in Escherichia coli strain DH5alpha
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Geobacillus kaustophilis RLPalpha-1 sequence determination and analysis, and phylogenetic analysis
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Microcystis aeruginosa RLPalpha-1 sequence determination and analysis, and phylogenetic analysis
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overexpression in Escherichia coli
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overexpression in Klebsiella pneumoniae strain CG253 leading to 5-6fold increased activity compared to the parent strain
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Rhodopseudomonas palustris RLPalpha-1 sequence determination and analysis, and phylogenetic analysis
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Rhodospirillum rubrum RLPalpha-1 sequence determination and analysis, and phylogenetic analysis
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
K122E
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the mutant fails to form a catalytic dimer and does not show any DK-MTP-1-P enolase activity
K122M
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the mutant fails to form a catalytic dimer and does not show any DK-MTP-1-P enolase activity
K122R
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the mutant has specific activity of 32% less than that of the wild type enzyme and a lower thermal stability than the wild type enzyme
K147A
site-directed mutagenesis, the mutant requires a 10fold greater concentration of protein for enolization of the natural substrate, reduced activity compared to the wild-type enzyme
K173A
site-directed mutagenesis, the mutant is able to catalyze enolization at approximately the same rate as the wild-type enzyme
K98A
site-directed mutagenesis, the mutant is unable to catalyze the enolase reaction
additional information
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a growth-defective mutant, in which the gene for the RLP is disrupted, is rescued by the gene for RuBisCO from the photosynthetic bacterium Rhodospirillum rubrum
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