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| Predicate | Object |
|---|---|
| rdf:type | |
| lifeskim:mentions | |
| pubmed:issue |
51
|
| pubmed:dateCreated |
1999-2-11
|
| pubmed:abstractText |
Epoxide hydrolase from Agrobacterium radiobacter AD1 catalyzes the enantioselective hydrolysis of styrene oxide with an E value of 16. The (R)-enantiomer of styrene oxide is first converted with a k(cat) of 3.8 s(-1), and the conversion of the (S)-enantiomer is inhibited. The latter is subsequently hydrolyzed with a k(cat) of 10.5 s(-1). The pre-steady-state kinetic parameters were determined for both enantiomers with stopped-flow fluorescence and rapid-quench techniques. For (R)-styrene oxide a four-step mechanism was needed to describe the data. It involved the formation of a Michaelis complex that is in rapid equilibrium with free enzyme and substrate, followed by rapid and reversible alkylation of the enzyme. A unimolecular isomerization of the alkylated enzyme precedes the hydrolysis of the covalent intermediate, which could be observed due to an enhancement of the intrinsic protein fluorescence during this step. The conversion of (S)-styrene oxide could be described by a three-step mechanism, which also involved reversible and rapid formation of an ester intermediate from a Michaelis complex and its subsequent slow hydrolysis as the rate-limiting step. The unimolecular isomerization step has not been observed for rat microsomal epoxide hydrolase, for which a kinetic mechanism was recently established [Tzeng, H.-F., Laughlin, L. T., Lin, S., and Armstrong, R. N. (1996) J. Am. Chem. Soc. 118, 9436-9437]. For both enantiomers of styrene oxide, the Km value was much lower than the substrate binding constant K(S) due to extensive accumulation of the covalent intermediate. The enantioselectivity was more pronounced in the alkylation rates than in the rate-limiting hydrolysis steps. The combined reaction schemes for (R)- and (S)-styrene oxide gave an accurate description of the epoxide hydrolase catalyzed kinetic resolution of racemic styrene oxide.
|
| pubmed:language |
eng
|
| pubmed:journal | |
| pubmed:citationSubset |
IM
|
| pubmed:chemical | |
| pubmed:status |
MEDLINE
|
| pubmed:month |
Dec
|
| pubmed:issn |
0006-2960
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| pubmed:author | |
| pubmed:issnType |
Print
|
| pubmed:day |
22
|
| pubmed:volume |
37
|
| pubmed:owner |
NLM
|
| pubmed:authorsComplete |
Y
|
| pubmed:pagination |
18119-27
|
| pubmed:dateRevised |
2009-11-19
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| pubmed:meshHeading |
pubmed-meshheading:9922181-Binding Sites,
pubmed-meshheading:9922181-Epoxide Hydrolases,
pubmed-meshheading:9922181-Epoxy Compounds,
pubmed-meshheading:9922181-Hydrolysis,
pubmed-meshheading:9922181-Isoenzymes,
pubmed-meshheading:9922181-Kinetics,
pubmed-meshheading:9922181-Protein Conformation,
pubmed-meshheading:9922181-Rhizobium,
pubmed-meshheading:9922181-Spectrometry, Fluorescence,
pubmed-meshheading:9922181-Stereoisomerism,
pubmed-meshheading:9922181-Substrate Specificity
|
| pubmed:year |
1998
|
| pubmed:articleTitle |
Kinetic mechanism of the enantioselective conversion of styrene oxide by epoxide hydrolase from Agrobacterium radiobacter AD1.
|
| pubmed:affiliation |
Department of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, The Netherlands.
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| pubmed:publicationType |
Journal Article
|