Information on EC 3.3.2.8 - limonene-1,2-epoxide hydrolase

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

EC NUMBER
COMMENTARY hide
3.3.2.8
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RECOMMENDED NAME
GeneOntology No.
limonene-1,2-epoxide hydrolase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
1,2-epoxymenth-8-ene + H2O = menth-8-ene-1,2-diol
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of epoxide
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Limonene and pinene degradation
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limonene degradation I (D-limonene)
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limonene degradation II (L-limonene)
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SYSTEMATIC NAME
IUBMB Comments
1,2-epoxymeth-8-ene hydrolase
Involved in the monoterpene degradation pathway of the actinomycete Rhodococcus erythropolis. The enzyme hydrolyses several alicyclic and 1-methyl-substituted epoxides, such as 1-methylcyclohexene oxide, indene oxide and cyclohexene oxide. It differs from the previously described epoxide hydrolases [EC 3.3.2.4 (trans-epoxysuccinate hydrolase), EC 3.3.2.6 (leukotriene-A4 hydrolase), EC 3.3.2.7 (hepoxilin-epoxide hydrolase), EC 3.3.2.9 (microsomal epoxide hydrolase) and EC 3.3.2.10 (soluble epoxide hydrolase)] as it is not inhibited by 2-bromo-4'-nitroacetophenone, diethyl dicarbonate, 4-fluorochalcone oxide or 1,10-phenanthroline. Both enantiomers of menth-8-ene-1,2-diol [i.e. (1R,2R,4S)-menth-8-ene-1,2-diol and (1S,2S,4R)-menth-8-ene-1,2-diol] are metabolized.
CAS REGISTRY NUMBER
COMMENTARY hide
216503-88-7
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
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Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
(1R,2S)-1-methylcyclohexane oxide + H2O
(1S,2S)-1-methylcyclohexane-1,2-diol
show the reaction diagram
(1R,2S,4R)-limonene-1,2-epoxide + H2O
(1S,2S,4R)-limonene-1,2-diol
show the reaction diagram
(1R,2S,4S)-limonene-1,2-epoxide + H2O
(1R,2R,4S)-limonene-1,2-diol
show the reaction diagram
(1S,2R)-1-methylcyclohexane oxide + H2O
(1R,2R)-1-methylcyclohexane-1,2-diol
show the reaction diagram
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-
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ir
(1S,2R,4R)-limonene-1,2-epoxide + H2O
(1S,2S,4R)-limonene-1,2-diol
show the reaction diagram
(1S,2R,4S)-limonene-1,2-epoxide + H2O
(1R,2R,4S)-limonene-1,2-diol
show the reaction diagram
the reaction mechanism involves epoxide protonation by Asp109, nucleophilic attack by water, and abstraction of a proton from water by Asp132. The isopropenyl group plays a crucial role because it restricts the half-chair conformation to one of the two possible helicities. In this conformation, attack on the different epoxide carbons will lead to either a chair-like or a twist-boat transition state structure, the latter resulting in a higher barrier. The regioselectivity is thus governed by conformational and not electronic factors
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ir
(1S2R4S)-limonene-1,2-epoxide + H2O
(1R,2R,4S)-limonene-1,2-diol
show the reaction diagram
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optically pure, diastereomeric excess above 99%
?
1,2-epoxy-2,6-dimethyl-5-heptane + H2O
(2S)-2,6-dimethyl-5-heptene-1,2-diol
show the reaction diagram
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?
1,2-epoxy-2-methyl-6-heptene + H2O
(2S)-2-methyl-6-heptene-1,2-diol
show the reaction diagram
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?
1,2-epoxy-2-methylheptane + H2O
(2S)-2-methylheptane-1,2-diol
show the reaction diagram
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?
1,2-epoxy-3-benzyl-2-methylpropane + H2O
(2S)-3-benzyl-2-methylpropane-1,2-diol
show the reaction diagram
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?
1-methylcyclohexene oxide + H2O
(1S,2S)-1-methylcyclohexane-1,2-diol
show the reaction diagram
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?
cyclohexene oxide + H2O
(1S,2S)-trans cyclohexanediol + (1R,2R)-trans-cyclohexanediol
show the reaction diagram
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?
cyclopentene-oxide + H2O
(R,R)-cyclopentene-1,2-diol + (S,S)-cyclopentene-1,2-diol
show the reaction diagram
indene oxide + H2O
indane-1,2-diol
show the reaction diagram
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?
limonene-1,2-eopxide + H2O
limonene-1,2-diol
show the reaction diagram
limonene-1,2-epoxide + H2O
limonene-1,2-diol
show the reaction diagram
rac-1-methyl-7-oxabicyclo[4.1.0]heptane + H2O
(1S,2S)-1-methylcyclohexane-1,2-diol + (1R,6S)-1-methyl-7-oxabicyclo[4.1.0]heptane
show the reaction diagram
rac-2-(phenoxymethyl)oxirane + H2O
(2S)-2-(phenoxymethyl)oxirane + (2R)-3-phenoxypropane-1,2-diol
show the reaction diagram
S-(-)-limonene + H2O
limonene-1,2-diol
show the reaction diagram
styrene-7,8-oxide + H2O
styrene-7,8-diol
show the reaction diagram
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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
(1R,2S,4R)-limonene-1,2-epoxide + H2O
(1S,2S,4R)-limonene-1,2-diol
show the reaction diagram
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optically pure, diastereomeric excess above 99%
?
(1R,2S,4S)-limonene-1,2-epoxide + H2O
(1R,2R,4S)-limonene-1,2-diol
show the reaction diagram
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optically pure, diastereomeric excess above 99%
?
(1S,2R,4R)-limonene-1,2-epoxide + H2O
(1S,2S,4R)-limonene-1,2-diol
show the reaction diagram
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optically pure, diastereomeric excess above 99%
?
(1S2R4S)-limonene-1,2-epoxide + H2O
(1R,2R,4S)-limonene-1,2-diol
show the reaction diagram
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optically pure, diastereomeric excess above 99%
?
limonene-1,2-eopxide + H2O
limonene-1,2-diol
show the reaction diagram
limonene-1,2-epoxide + H2O
limonene-1,2-diol
show the reaction diagram
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
hexanamide
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competitive
Hexylamine
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competitive
valpromide
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i.e. dipropylacetamide, competitive
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.4 - 3.7
styrene-7,8-oxide
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.002 - 0.47
styrene-7,8-oxide
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2
hexanamide
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pH 7.4, 1% ethanol in final assay mixture
0.035
Hexylamine
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pH 7.4, 1% ethanol in final assay mixture
0.1
valpromide
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pH 7.4, 1% ethanol in final assay mixture
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
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very broad pH optimum
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 10.5
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below pH 5 chemical hydrolysis overtook biological hydrolysis rate
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 45
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rapid inactivation above 45°C
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
15000
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gel filtration
16520
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deduced from amino acid sequence
17000
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SDS-PAGE
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
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crystallization data
monomer
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gel filtration
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
modeling of substrate 1,2-epoxymenth-8-ene into the active site of crystal structure and evaluation of the roles of residues Arg99, Tyr53 and Asn55 by QM/MM-scannedenergy mapping
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selenomethionine-substituted enzyme
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
49
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15 min, 50% resiudal activity for wild-type
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
gel filtration, hydroxyapatite, anion exchange
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D101A
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catalytically inactive, D101 is the acid catalyst that protonates the epoxide oxygen
D101N
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catalytically inactive, D101 is the acid catalyst that protonates the epoxide oxygen
L114C/I116V
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substrate cyclopentene-oxide, 72% conversion, (S,S)-product with 68% enantiomeric excess
L114I/I116V
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substrate cyclopentene-oxide, 74% conversion, (S,S)-product with 50% enantiomeric excess
L114V/I116V
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substrate cyclopentene-oxide, 72% conversion, (S,S)-product with 60% enantiomeric excess
L74I/I80C
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substrate cyclopentene-oxide, 75% conversion, (R,R)-product with 66% enantiomeric excess
L74I/I80V
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substrate cyclopentene-oxide, 75% conversion, (R,R)-product with 58% enantiomeric excess
L74V/I80V
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substrate cyclopentene-oxide, 67% conversion, (R,R)-product with 53% enantiomeric excess
M32C/I80F/L114C/I116V
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substrate rac-2-(phenoxymethyl)oxirane, 31% conversion, (2R)-product with 92% enantiomeric excess. Substrate rac-1-methyl-7-oxabicyclo[4.1.0]heptane, 99% conversion, (1S,2S)-product with 55% enantiomeric excess
M32L/L35C
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substrate cyclopentene-oxide, 78% conversion, (S,S)-product with 16% enantiomeric excess
M32L/L35F
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substrate cyclopentene-oxide, 79% conversion, (S,S)-product with 24% enantiomeric excess
M32L/L35V
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substrate cyclopentene-oxide, 78% conversion, (S,S)-product with 10% enantiomeric excess
M78F/V83I
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substrate cyclopentene-oxide, 82% conversion, (R,R)-product with 29% enantiomeric excess
M78I/V83I
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substrate cyclopentene-oxide, 80% conversion, (R,R)-product with 13% enantiomeric excess
M78V/V83I
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substrate cyclopentene-oxide, 68% conversion, (R,R)-product with 7% enantiomeric excess
N55A
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little residual activity
N55D
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catalytically inactive
N55D/D132N
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catalytically inactive, folding correct
R99A
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catalytically inactive
R99H
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catalytically inactive
R99K
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catalytically inactive
R99Q
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catalytically inactive
Y53F
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partially active
L74V/I80V
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substrate cyclopentene-oxide, 67% conversion, (R,R)-product with 53% enantiomeric excess
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M32L/L35C
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substrate cyclopentene-oxide, 78% conversion, (S,S)-product with 16% enantiomeric excess
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M32L/L35F
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substrate cyclopentene-oxide, 79% conversion, (S,S)-product with 24% enantiomeric excess
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M32L/L35V
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substrate cyclopentene-oxide, 78% conversion, (S,S)-product with 10% enantiomeric excess
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additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
synthesis