Information on EC 5.4.99.14 - 4-Carboxymethyl-4-methylbutenolide mutase

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The expected taxonomic range for this enzyme is: Bacteria

EC NUMBER
COMMENTARY hide
5.4.99.14
-
RECOMMENDED NAME
GeneOntology No.
4-Carboxymethyl-4-methylbutenolide mutase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
4-Carboxymethyl-4-methylbut-2-en-1,4-olide = 4-carboxymethyl-3-methylbut-2-en-1,4-olide
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
intramolecular transfer reaction
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-
-
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
4-methylcatechol degradation (ortho cleavage)
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Microbial metabolism in diverse environments
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Toluene degradation
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SYSTEMATIC NAME
IUBMB Comments
4-Carboxymethyl-4-methylbut-2-en-1,4-olide methylmutase
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CAS REGISTRY NUMBER
COMMENTARY hide
115300-03-3
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
gene mmlI
UniProt
Manually annotated by BRENDA team
N657
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-
Manually annotated by BRENDA team
N657
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-
Manually annotated by BRENDA team
N5
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-
Manually annotated by BRENDA team
N75
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-
Manually annotated by BRENDA team
N75
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Manually annotated by BRENDA team
N75
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Manually annotated by BRENDA team
strains BCN1, BCN2 and 4PH1
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-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
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
(4R,5R)-5-chloro-3-methylmuconolactone
trans-3-methyldienelactone
show the reaction diagram
(4S)-4-methylmuconolactone
(4S)-3-methylmuconolactone
show the reaction diagram
(4S,5S)-5-chloro-3-methylmuconolactone
trans-3-methyldienelactone
show the reaction diagram
1-methylbis-lactone
?
show the reaction diagram
1-Methylbislactone
4-Carboxymethyl-3-methylbut-2-en-1,4-olide
show the reaction diagram
3-methylmuconolactone
4-methyl-3-oxoadipate enollactone
show the reaction diagram
4-Carboxymethyl-4-methylbut-2-en-1,4-olide
4-Carboxymethyl-3-methylbut-2-en-1,4-olide
show the reaction diagram
4-Carboxymethyl-4-methylbut-2-en-1,4-olide
?
show the reaction diagram
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enzyme of the modified 3-oxoadipate pathway in the Gram-negative bacterium Alcaligenes eutrophus JMP 134
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-
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4-methyl-muconolactone
3-methyl-muconolactone
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
4-Carboxymethyl-4-methylbut-2-en-1,4-olide
?
show the reaction diagram
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enzyme of the modified 3-oxoadipate pathway in the Gram-negative bacterium Alcaligenes eutrophus JMP 134
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-
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4-methyl-muconolactone
3-methyl-muconolactone
show the reaction diagram
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
(+)-muconolactone
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(-)1-Methyl-8-bromobislactone
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weak
2,4-Dimethyl-2-enelactone
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weak
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2-Bromo-4-methylbutan-1,4-olide
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weak
2-Bromobutan-1,4-olide
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2-Methyl-2-enelactone
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weak
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2-Methylbutan-1,4-olide
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3-Methyl-2-enelactone
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weak
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4-carboxymethylbut-2-en-4-olide
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4-chloromercuribenzoate
4-methyl-2-enelactone
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weak
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4-Methylbut-3-en-1,4-olide
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angelicalactone
4-methylbutan-1,4-olide
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trans-dienelactone
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weak
additional information
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.045 - 0.4583
(4S)-4-methylmuconolactone
0.028
1-methyl-3,7-dioxo-2,6-dioxabicyclo[3.3.0]octane
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0.17
1-methyl-bislactone
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0.176 - 0.23
4-carboxymethyl-4-methylbut-2-en-1,4-olide
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
5.8 - 47.2
(4S)-4-methylmuconolactone
3.75
1-methyl-3,7-dioxo-2,6-dioxabicyclo[3.3.0]octane
Cupriavidus necator
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98.5
4-carboxymethyl-4-methylbut-2-en-1,4-olide
Cupriavidus necator
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kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
100 - 500
(4S)-4-methylmuconolactone
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
28.8
toward 1-methylbis-lactone, in phosphate buffer, 50 mM, pH 7.3
57.3
toward (4S)-4-methylmuconolactone, in phosphate buffer, 50 mM, pH 7.3
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 8.5
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active in this range, rapid decrease of activity above pH 8.5
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25000
gel filtration
75500
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Rhodococcus rhodochrous, gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homodimer
monomer
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1 * 40000, Alcaligenes eutrophus (JMP 134, cloned into Pseudomonas sp. B13 FR1), SDS-PAGE
tetramer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
in complex with (4S)-4-methylmuconolactone and (4S)-3-methylmuconolactone, sitting drop vapor diffusion method, at 19C, using 25% (w/v) polyethylene glycol 1500, 0.1 M MMT buffer, pH 5 (DL-malic acid, MES, and Tris base in a 1:2:2 molar ratio)
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
Chelating agents like EDTA or thiol-group modifying reagents like DTT stabilize
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-30C, slow loss of activity on prolonged storage
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4C or -20C, highly purified enzyme is rather unstable
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4C, 24 h, 10% loss of activity, highly purified enzyme, 5% loss of activity with addition of chelating agents like EDTA or thiol-group modifying reagents like DTT
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
JMP 134, cloned into Pseudomonas sp. B13 FR1
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MonoQ column chromatography
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Strep-Tactin-Sepharose column chromatography
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli JM109 cells
gene mmlI, DNA and amino acid sequence determination and analysis, expression in Escherichia coli strain BL21(DE3)
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C67S
incubation of variant C67S with 4-chloromercuribenzoate does not significantly decrease its activity (above 65% of activity remaining after a 30-min incubation)
H26A
the mutant shows a marked decrease in catalytic activity (kcat) by two orders of magnitude
H52A
the mutation results in an increase in Km by a factor of 9, but only a small change in kcat
Y39F
the mutant displays reduced activity compared to the wild type enzyme
C67S
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incubation of variant C67S with 4-chloromercuribenzoate does not significantly decrease its activity (above 65% of activity remaining after a 30-min incubation)
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H26A
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the mutant shows a marked decrease in catalytic activity (kcat) by two orders of magnitude
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H52A
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the mutation results in an increase in Km by a factor of 9, but only a small change in kcat
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Y39F
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the mutant displays reduced activity compared to the wild type enzyme
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