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ABSTRACT: The presence of different hydrophilic organic solvents or a water soluble polymer such as PEG 4000 led to an enhancement in the enzymatic activity of the M446G mutant of phenylacetone monooxygenase when it is employed in enantioselective sulfoxidations and Baeyer-Villiger reactions. By solvent engineering new substrates were found to be effectively converted by this Baeyer-Villiger monooxygenase. The use of 5% methanol together with the weak anion exchange resin Lewatit MP62 also allows the dynamic kinetic resolution of a set of racemic benzylketones. By this approach (S)-benzylesters could be obtained with high yields and optical purities.
Enzyme and microbial technology. 01/2012; 50(1):43-9.
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ABSTRACT: BVMOs make a play: The dynamic kinetic resolution of racemic α-alkyl-β-ketoesters was performed through a selective Baeyer-Villiger oxidation employing different Baeyer-Villiger monooxygenases (BVMOs) in mild basic media. The product diesters were obtained with excellent yields and enantioselectivities, and used as precursors for optically active α-hydroxy esters.
Angewandte Chemie International Edition 07/2011; 50(36):8387-90. · 13.45 Impact Factor
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ABSTRACT: Type I Baeyer-Villiger monooxygenases (BVMOs) strongly prefer NADPH over NADH as an electron donor. In order to elucidate the molecular basis for this coenzyme specificity, we have performed a site-directed mutagenesis study on phenylacetone monooxygenase (PAMO) from Thermobifida fusca. Using sequence alignments of type I BVMOs and crystal structures of PAMO and cyclohexanone monooxygenase in complex with NADP(+), we identified four residues that could interact with the 2'-phosphate moiety of NADPH in PAMO. The mutagenesis study revealed that the conserved R217 is essential for binding the adenine moiety of the nicotinamide coenzyme while it also contributes to the recognition of the 2'-phosphate moiety of NADPH. The substitution of T218 did not have a strong effect on the coenzyme specificity. The H220N and H220Q mutants exhibited a ~3-fold improvement in the catalytic efficiency with NADH while the catalytic efficiency with NADPH was hardly affected. Mutating K336 did not increase the activity of PAMO with NADH, but it had a significant and beneficial effect on the enantioselectivity of Baeyer-Villiger oxidations and sulfoxidations. In conclusion, our results indicate that the function of NADPH in catalysis cannot be easily replaced by NADH. This finding is in line with the complex catalytic mechanism and the vital role of the coenzyme in BVMOs.
Applied Microbiology and Biotechnology 11/2010; 88(5):1135-43. · 3.42 Impact Factor
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ABSTRACT: 4-Hydroxyacetophenone monooxygenase from Pseudomonas fluorescens ACB was employed in the presence of a weak anion exchange resin to perform dynamic kinetic resolutions of racemic benzyl ketones with high conversions and good optical purities. Different parameters that affect to the efficiency of the enzymatic Baeyer-Villiger oxidation and racemisation were analyzed in order to optimize the activity and selectivity of the biocatalytic system.
Organic & Biomolecular Chemistry 03/2010; 8(5):1121-5. · 3.70 Impact Factor
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ABSTRACT: Parallel interconnected kinetic asymmetric transformations were performed in order to obtain enantioenriched derivatives starting from a set of racemic or prochiral compounds. Thus, in a one-pot reaction using two redox biocatalysts (a BVMO and an ADH) and a catalytic amount of cofactor that acts as a mediator, enantioenriched ketones, sulfoxides, and sec-alcohols were concurrently obtained in a strict parallel way, minimising the quantity of reagents employed. By selecting the appropriate biocatalysts, this methodology represents a potential tool for performing stereodivergent transformations.
Organic & Biomolecular Chemistry 03/2010; 8(6):1431-7. · 3.70 Impact Factor
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ABSTRACT: The application of three BVMOs for the enantioselective oxidation of 3-phenylbutan-2-ones with different substituents in the aromatic moiety is described. By choosing the appropriate biocatalyst and substrate combination, chiral ketones and esters can be obtained with excellent enantiopurities. This methodology could also be applied to the resolution of racemic α-alkyl benzylketones with longer alkyl chains as well as with two substituted α-substituted benzylacetones. A kinetic analysis revealed that the BVMOs studied effectively convert all tested compounds showing that the enzymes are tolerant towards the substrate structure while being highly enantioselective. These properties render BVMOs as valuable biocatalysts for the preparation of compounds with high interest in organic synthesis.
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ABSTRACT: The present study describes the first-time usage of an isolated thermostable Baeyer–Villiger monooxygenase (phenylacetone monooxygenase, PAMO) in the presence of ionic liquids. The stability, activity and selectivity of PAMO as an oxidative enzyme in the presence of different ionic liquids were studied. This revealed that the addition of some specific ionic liquids, such AmmoengTM 102 and [bmim]MeSO4, can significantly enhance the E-value in the oxidation of racemic benzylketones. Moreover, the use of ionic liquids increases the optimal substrate concentration for performing Baeyer–Villiger oxidation, thereby extending the biocatalytic repertoire of PAMO for synthetic applications.
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ABSTRACT: The enantioselective kinetic resolution of a set of racemic substituted 3-phenylbutan-2-ones employing phenylacetone monooxygenase (PAMO) in non-conventional media was performed. The studies have revealed the effects of a range of solvents on the biocatalytic properties of the biocatalyst. Also, the enzymatic oxidation of α-acetylphenylacetonitrile was performed using organic cosolvents. This has resulted in a dynamic kinetic resolution of this cyanoketone yielding enantiopure (R)-2-acetoxyphenylacetonitrile with moderate yields depending on the reaction conditions employed.
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ABSTRACT: A set of racemic cyclic and linear ketones, as well as 2-phenylpropionaldehyde, were tested as substrates in the enzymatic Baeyer–Villiger oxidation catalyzed by two Baeyer–Villiger monooxygenases: phenylacetone monooxygenase (PAMO) and 4-hydroxyacetophenone monooxygenase (HAPMO). Excellent enantioselectivites (E > 200) can be obtained in the kinetic resolution processes depending on the substrate structure and the reaction conditions. The parameters affecting the biocatalytic properties of these enzymes were also studied, in order to establish a deeper understanding of these novel biocatalysts.
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ABSTRACT: Phenylacetone monooxygense (PAMO) from Thermobifida fusca was employed for the asymmetric oxidation of thioanisole (sulfooxidation) and of racemic 2-phenylpropionaldehyde (Baeyer–Villiger oxidation). A pH dependence of enantioselectivity was observed in both cases. Two different residues, with pKa values of 7.8±0.2 and 9.2±0.2, appeared to be responsible for the pH effects on PAMO enantioselectivity. The protonation of Arg337 and the FAD:C4a-hydroperoxide/FAD:C4a-peroxide equilibrium were identified as the major factors responsible for the fine-tuning of PAMO enantioselectivity in Baeyer–Villiger oxidation and sulfooxidation, respectively.
