BRENDA - Enzyme Database
show all sequences of 3.3.2.8

Manipulating the stereoselectivity of limonene epoxide hydrolase by directed evolution based on iterative saturation mutagenesis

Zheng, H.; Reetz, M.T.; J. Am. Chem. Soc. 132, 15744-15751 (2010)

Data extracted from this reference:

Application
Application
Commentary
Organism
synthesis
application of directed evolution using iterative saturation mutagenesis as a means to engineer LEH mutants showing broad substrate scope with high stereoselectivity. Mutants are obtained which catalyze the desymmetrization of cyclopentene-oxide with stereoselective formation of either the (R,R)- or the (S,S)-diol on an optional basis. The mutants prove to be excellent catalysts for the desymmetrization of other meso-epoxides and for the hydrolytic kinetic resolution of racemic substrates
Rhodococcus erythropolis
Engineering
Protein Variants
Commentary
Organism
L114C/I116V
substrate cyclopentene-oxide, 72% conversion, (S,S)-product with 68% enantiomeric excess
Rhodococcus erythropolis
L114I/I116V
substrate cyclopentene-oxide, 74% conversion, (S,S)-product with 50% enantiomeric excess
Rhodococcus erythropolis
L114V/I116V
substrate cyclopentene-oxide, 72% conversion, (S,S)-product with 60% enantiomeric excess
Rhodococcus erythropolis
L74I/I80C
substrate cyclopentene-oxide, 75% conversion, (R,R)-product with 66% enantiomeric excess
Rhodococcus erythropolis
L74I/I80V
substrate cyclopentene-oxide, 75% conversion, (R,R)-product with 58% enantiomeric excess
Rhodococcus erythropolis
L74V/I80V
substrate cyclopentene-oxide, 67% conversion, (R,R)-product with 53% enantiomeric excess
Rhodococcus erythropolis
M32C/I80F/L114C/I116V
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
Rhodococcus erythropolis
M32L/L35C
substrate cyclopentene-oxide, 78% conversion, (S,S)-product with 16% enantiomeric excess
Rhodococcus erythropolis
M32L/L35F
substrate cyclopentene-oxide, 79% conversion, (S,S)-product with 24% enantiomeric excess
Rhodococcus erythropolis
M32L/L35V
substrate cyclopentene-oxide, 78% conversion, (S,S)-product with 10% enantiomeric excess
Rhodococcus erythropolis
M78F/V83I
substrate cyclopentene-oxide, 82% conversion, (R,R)-product with 29% enantiomeric excess
Rhodococcus erythropolis
M78I/V83I
substrate cyclopentene-oxide, 80% conversion, (R,R)-product with 13% enantiomeric excess
Rhodococcus erythropolis
M78V/V83I
substrate cyclopentene-oxide, 68% conversion, (R,R)-product with 7% enantiomeric excess
Rhodococcus erythropolis
additional information
application of directed evolution using iterative saturation mutagenesis as a means to engineer LEH mutants showing broad substrate scope with high stereoselectivity. Mutants are obtained which catalyze the desymmetrization of cyclopentene-oxide with stereoselective formation of either the (R,R)- or the (S,S)-diol on an optional basis. The mutants prove to be excellent catalysts for the desymmetrization of other meso-epoxides and for the hydrolytic kinetic resolution of racemic substrates
Rhodococcus erythropolis
Organism
Organism
UniProt
Commentary
Textmining
Rhodococcus erythropolis
-
-
-
Rhodococcus erythropolis DCL 14
-
-
-
Substrates and Products (Substrate)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
Substrate Product ID
cyclopentene-oxide + H2O
-
715269
Rhodococcus erythropolis
(R,R)-cyclopentene-1,2-diol + (S,S)-cyclopentene-1,2-diol
wild-type, 72% conversion, (R,R)-product with 14% enantiomeric excess
-
-
?
cyclopentene-oxide + H2O
-
715269
Rhodococcus erythropolis DCL 14
(R,R)-cyclopentene-1,2-diol + (S,S)-cyclopentene-1,2-diol
wild-type, 72% conversion, (R,R)-product with 14% enantiomeric excess
-
-
?
rac-1-methyl-7-oxabicyclo[4.1.0]heptane + H2O
-
715269
Rhodococcus erythropolis
(1S,2S)-1-methylcyclohexane-1,2-diol + (1R,6S)-1-methyl-7-oxabicyclo[4.1.0]heptane
wild-type, 99% conversion, 19% enantiomeric excess for (1S,2S)-product
-
-
-
rac-1-methyl-7-oxabicyclo[4.1.0]heptane + H2O
-
715269
Rhodococcus erythropolis DCL 14
(1S,2S)-1-methylcyclohexane-1,2-diol + (1R,6S)-1-methyl-7-oxabicyclo[4.1.0]heptane
wild-type, 99% conversion, 19% enantiomeric excess for (1S,2S)-product
-
-
-
rac-2-(phenoxymethyl)oxirane + H2O
-
715269
Rhodococcus erythropolis
(2S)-2-(phenoxymethyl)oxirane + (2R)-3-phenoxypropane-1,2-diol
wild-type, 33% conversion, 37% enantiomeric excess
-
-
?
rac-2-(phenoxymethyl)oxirane + H2O
-
715269
Rhodococcus erythropolis DCL 14
(2S)-2-(phenoxymethyl)oxirane + (2R)-3-phenoxypropane-1,2-diol
wild-type, 33% conversion, 37% enantiomeric excess
-
-
?
Temperature Stability [°C]
Temperature Stability Minimum [°C]
Temperature Stability Maximum [°C]
Commentary
Organism
49
-
15 min, 50% resiudal activity for wild-type
Rhodococcus erythropolis
Application (protein specific)
Application
Commentary
Organism
synthesis
application of directed evolution using iterative saturation mutagenesis as a means to engineer LEH mutants showing broad substrate scope with high stereoselectivity. Mutants are obtained which catalyze the desymmetrization of cyclopentene-oxide with stereoselective formation of either the (R,R)- or the (S,S)-diol on an optional basis. The mutants prove to be excellent catalysts for the desymmetrization of other meso-epoxides and for the hydrolytic kinetic resolution of racemic substrates
Rhodococcus erythropolis
Engineering (protein specific)
Protein Variants
Commentary
Organism
L114C/I116V
substrate cyclopentene-oxide, 72% conversion, (S,S)-product with 68% enantiomeric excess
Rhodococcus erythropolis
L114I/I116V
substrate cyclopentene-oxide, 74% conversion, (S,S)-product with 50% enantiomeric excess
Rhodococcus erythropolis
L114V/I116V
substrate cyclopentene-oxide, 72% conversion, (S,S)-product with 60% enantiomeric excess
Rhodococcus erythropolis
L74I/I80C
substrate cyclopentene-oxide, 75% conversion, (R,R)-product with 66% enantiomeric excess
Rhodococcus erythropolis
L74I/I80V
substrate cyclopentene-oxide, 75% conversion, (R,R)-product with 58% enantiomeric excess
Rhodococcus erythropolis
L74V/I80V
substrate cyclopentene-oxide, 67% conversion, (R,R)-product with 53% enantiomeric excess
Rhodococcus erythropolis
M32C/I80F/L114C/I116V
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
Rhodococcus erythropolis
M32L/L35C
substrate cyclopentene-oxide, 78% conversion, (S,S)-product with 16% enantiomeric excess
Rhodococcus erythropolis
M32L/L35F
substrate cyclopentene-oxide, 79% conversion, (S,S)-product with 24% enantiomeric excess
Rhodococcus erythropolis
M32L/L35V
substrate cyclopentene-oxide, 78% conversion, (S,S)-product with 10% enantiomeric excess
Rhodococcus erythropolis
M78F/V83I
substrate cyclopentene-oxide, 82% conversion, (R,R)-product with 29% enantiomeric excess
Rhodococcus erythropolis
M78I/V83I
substrate cyclopentene-oxide, 80% conversion, (R,R)-product with 13% enantiomeric excess
Rhodococcus erythropolis
M78V/V83I
substrate cyclopentene-oxide, 68% conversion, (R,R)-product with 7% enantiomeric excess
Rhodococcus erythropolis
additional information
application of directed evolution using iterative saturation mutagenesis as a means to engineer LEH mutants showing broad substrate scope with high stereoselectivity. Mutants are obtained which catalyze the desymmetrization of cyclopentene-oxide with stereoselective formation of either the (R,R)- or the (S,S)-diol on an optional basis. The mutants prove to be excellent catalysts for the desymmetrization of other meso-epoxides and for the hydrolytic kinetic resolution of racemic substrates
Rhodococcus erythropolis
Substrates and Products (Substrate) (protein specific)
Substrates
Commentary Substrates
Literature (Substrates)
Organism
Products
Commentary (Products)
Literature (Products)
Organism (Products)
Reversibility
ID
cyclopentene-oxide + H2O
-
715269
Rhodococcus erythropolis
(R,R)-cyclopentene-1,2-diol + (S,S)-cyclopentene-1,2-diol
wild-type, 72% conversion, (R,R)-product with 14% enantiomeric excess
-
-
?
cyclopentene-oxide + H2O
-
715269
Rhodococcus erythropolis DCL 14
(R,R)-cyclopentene-1,2-diol + (S,S)-cyclopentene-1,2-diol
wild-type, 72% conversion, (R,R)-product with 14% enantiomeric excess
-
-
?
rac-1-methyl-7-oxabicyclo[4.1.0]heptane + H2O
-
715269
Rhodococcus erythropolis
(1S,2S)-1-methylcyclohexane-1,2-diol + (1R,6S)-1-methyl-7-oxabicyclo[4.1.0]heptane
wild-type, 99% conversion, 19% enantiomeric excess for (1S,2S)-product
-
-
-
rac-1-methyl-7-oxabicyclo[4.1.0]heptane + H2O
-
715269
Rhodococcus erythropolis DCL 14
(1S,2S)-1-methylcyclohexane-1,2-diol + (1R,6S)-1-methyl-7-oxabicyclo[4.1.0]heptane
wild-type, 99% conversion, 19% enantiomeric excess for (1S,2S)-product
-
-
-
rac-2-(phenoxymethyl)oxirane + H2O
-
715269
Rhodococcus erythropolis
(2S)-2-(phenoxymethyl)oxirane + (2R)-3-phenoxypropane-1,2-diol
wild-type, 33% conversion, 37% enantiomeric excess
-
-
?
rac-2-(phenoxymethyl)oxirane + H2O
-
715269
Rhodococcus erythropolis DCL 14
(2S)-2-(phenoxymethyl)oxirane + (2R)-3-phenoxypropane-1,2-diol
wild-type, 33% conversion, 37% enantiomeric excess
-
-
?
Temperature Stability [°C] (protein specific)
Temperature Stability Minimum [°C]
Temperature Stability Maximum [°C]
Commentary
Organism
49
-
15 min, 50% resiudal activity for wild-type
Rhodococcus erythropolis
Other publictions for EC 3.3.2.8
No.
1st author
Pub Med
title
organims
journal
volume
pages
year
Activating Compound
Application
Cloned(Commentary)
Crystallization (Commentary)
Engineering
General Stability
Inhibitors
KM Value [mM]
Localization
Metals/Ions
Molecular Weight [Da]
Natural Substrates/ Products (Substrates)
Organic Solvent Stability
Organism
Oxidation Stability
Posttranslational Modification
Purification (Commentary)
Reaction
Renatured (Commentary)
Source Tissue
Specific Activity [micromol/min/mg]
Storage Stability
Substrates and Products (Substrate)
Subunits
Synonyms
Temperature Optimum [°C]
Temperature Range [°C]
Temperature Stability [°C]
Turnover Number [1/s]
pH Optimum
pH Range
pH Stability
Cofactor
Ki Value [mM]
pI Value
IC50 Value
Activating Compound (protein specific)
Application (protein specific)
Cloned(Commentary) (protein specific)
Cofactor (protein specific)
Crystallization (Commentary) (protein specific)
Engineering (protein specific)
General Stability (protein specific)
IC50 Value (protein specific)
Inhibitors (protein specific)
Ki Value [mM] (protein specific)
KM Value [mM] (protein specific)
Localization (protein specific)
Metals/Ions (protein specific)
Molecular Weight [Da] (protein specific)
Natural Substrates/ Products (Substrates) (protein specific)
Organic Solvent Stability (protein specific)
Oxidation Stability (protein specific)
Posttranslational Modification (protein specific)
Purification (Commentary) (protein specific)
Renatured (Commentary) (protein specific)
Source Tissue (protein specific)
Specific Activity [micromol/min/mg] (protein specific)
Storage Stability (protein specific)
Substrates and Products (Substrate) (protein specific)
Subunits (protein specific)
Temperature Optimum [°C] (protein specific)
Temperature Range [°C] (protein specific)
Temperature Stability [°C] (protein specific)
Turnover Number [1/s] (protein specific)
pH Optimum (protein specific)
pH Range (protein specific)
pH Stability (protein specific)
pI Value (protein specific)
Expression
General Information
General Information (protein specific)
Expression (protein specific)
KCat/KM [mM/s]
KCat/KM [mM/s] (protein specific)
752350
Rinaldi
-
Understanding complex mechani ...
Rhodococcus erythropolis, Rhodococcus erythropolis DCL14
ACS Catal.
8
5698-5707
2018
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-
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2
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7
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1
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1
1
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4
1
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-
2
2
-
-
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753976
Sun
Structural and computational ...
uncultured organism
J. Am. Chem. Soc.
140
310-318
2018
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-
1
7
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-
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1
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5
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2
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1
7
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5
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1
1
-
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752348
Li
-
Multiparameter optimization i ...
Rhodococcus erythropolis, Rhodococcus erythropolis DCL14
ACS Catal.
6
3679-3687
2016
-
-
1
1
15
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-
11
-
-
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7
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1
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4
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2
1
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2
9
1
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1
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1
15
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11
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1
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4
-
1
-
2
9
1
-
-
-
-
-
-
-
11
11
731857
Molina
Comparative study of the bioco ...
Fusarium oxysporum 152B, Fusarium oxysporum
Food Chem.
174
606-613
2015
-
-
-
-
-
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3
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2
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1
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2
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-
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-
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-
753271
Ferrandi
-
Efficient epoxide hydrolase c ...
Rhodococcus erythropolis, Rhodococcus erythropolis DCL14, uncultured organism
ChemCatChem
7
3171-3178
2015
-
-
-
-
2
-
-
-
-
-
-
4
-
9
-
-
-
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8
-
13
-
9
3
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-
2
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-
3
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4
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8
-
13
-
3
-
-
-
3
-
-
-
-
-
-
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-
-
753544
Ferrandi
Discovery and characterizatio ...
Rhodococcus erythropolis, uncultured organism
FEBS J.
282
2879-2894
2015
-
-
1
1
2
-
3
-
-
-
-
3
-
4
-
-
1
2
-
-
-
-
33
2
9
3
-
3
-
3
-
-
-
-
-
-
-
-
2
-
2
2
-
-
3
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3
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2
-
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-
-
33
3
3
-
3
-
3
-
-
-
-
3
5
-
-
-
752797
Hou
QM/MM study of the mechanism ...
Rhodococcus erythropolis, Rhodococcus erythropolis DCL14
Biochim. Biophys. Acta
1824
263-268
2012
-
-
-
-
3
-
-
-
1
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2
-
7
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1
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10
1
3
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3
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1
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2
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10
1
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2
2
-
-
-
714359
Hou
QM/MM study of the mechanism o ...
Rhodococcus erythropolis, Rhodococcus erythropolis DCL14
Biochim. Biophys. Acta
1824
263-268
2011
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1
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2
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7
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1
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2
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2
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1
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2
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2
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-
-
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-
-
-
-
-
-
715269
Zheng
Manipulating the stereoselecti ...
Rhodococcus erythropolis, Rhodococcus erythropolis DCL 14
J. Am. Chem. Soc.
132
15744-15751
2010
-
1
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-
14
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3
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6
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1
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1
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14
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6
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1
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661940
Hopmann
Catalytic mechanism of limonen ...
Rhodococcus erythropolis, Rhodococcus erythropolis DCL14
J. Am. Chem. Soc.
127
14339-14347
2005
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8
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13
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655334
Arand
Structure of Rhodococcus eryth ...
Rhodococcus erythropolis
EMBO J.
22
2583-2592
2003
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1
10
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3
2
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2
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1
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1
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10
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3
3
2
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1
1
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3
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208936
Van der Werf
-
Substrate specificity and ster ...
Rhodococcus erythropolis
Appl. Microbiol. Biotechnol.
52
380-385
1999
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208937
Barbirato
The Rhodococcus erythropolis D ...
Rhodococcus erythropolis
FEBS Lett.
438
293-296
1998
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1
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208938
Van der Werf
Limonene-1,2-epoxide hydrolase ...
Rhodococcus erythropolis
J. Bacteriol.
180
5052-5057
1998
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2
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1
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