Blending Baeyer-Villiger monooxygenases: using a robust BVMO as a scaffold for creating chimeric enzymes with novel catalytic properties

The thermostable Baeyer-Villiger monooxygenase (BVMO) phenylacetone monooxygenase (PAMO) is used as a scaffold to introduce novel selectivities from other BVMOs or the metagenome by structure-inspired subdomain exchanges. This yields biocatalysts with new preferences in the oxidation of sulfides and the Baeyer-Villiger oxidation of ketones, all while maintaining most of the original thermostability.     

Radical-mediated thiodesulfonylation of the vinyl sulfones: access to (α-fluoro)vinyl sulfides

Radical-mediated thiodesulfonylation of the vinyl and (α-fluoro)vinyl sulfones, derived from aldehydes and ketones, with aryl thiols in organic or aqueous medium provided access to vinyl and (α-fluoro)vinyl sulfides. The vinyl sulfides were formed predominantly with E stereochemistry independent of the stereochemistry of the starting vinyl sulfones.     

4-Hydroxyacetophenone monooxygenase from Pseudomonas fluorescens ACB as an oxidative biocatalyst in the synthesis of optically active sulfoxides

Recombinant 4-hydroxyacetophenone monooxygenase (HAPMO) from Pseudomonas fluorescens ACB has been tested as a catalyst in sulfoxidation reactions on a set of aromatic sulfides. With a few exceptions, excellent enantioselectivities in the synthesis of chiral phenyl and benzyl sulfoxides were achieved. The bacterial Baeyer–Villiger monooxygenase was also shown to accept racemic sulfoxides, a prochiral diketone and an organoboron compound as substrates. This study demonstrates the great biocatalytic potential of this novel oxidative enzyme.     

Regio- and stereoselective preparation of highly substituted tertiary homoallylic alcohols

The titanocene(II)-promoted reaction of α-(benzyldimethylsilyl)allylic sulfides with ketones proceeded with high regio- and stereoselectivity to give δ-silylhomoallylic alcohols. The following palladium catalyzed cross-couplings with organic halides produced anti-(E)-β,δ-disubstituted tertiary homoallylic alcohols with complete retention of configuration.     

Simultaneous biocatalyst production and baeyer-villiger oxidation for bioonversion of cyclohexanone by recombinant Escherichia coli expressing cyclohexanone monooxygenase

Cyclohexanone monooxygenase (CHMO) catalyzing Baeyer-Villiger oxidation converts cyclic ketones into optically pure lactones, which have been used as building blocks in organic synthesis. A recombinant Escherichia coli BL21(DE3)/pMM4 expressing CHMO originated from Acinetobacter sp. NCIB 9871 was used to produce ε-caprolactone through a simultaneous biocatalyst production and Baeyer-Villiger oxidation (SPO) process. Afed-batch process was designed to obtain high cell density for improvin production of ε-caprolactone. The fed-batch SPO process have the best results, 10.2 g/L of ε-caprolactone and 0.34 g/(L·h) of productivity, corresponding to a 10.5- and 3.4-fold enhancement compared with those of the batch SPO, respectively.     

Baeyer-Villiger单加氧酶在有机合成中的应用

综述了Baeyer-Villiger单加氧酶在有机合成中的应用.较之传统的化学反应, 氧化酶催化剂反应有较好的选择性、可控性和经济性. 环己酮加氧酶是一种还原型辅酶I (NADPH)依赖型氧化酶, 是最早被报道能够催化Baeyer-Villiger氧化的酶. 这些重要反应产生了合成化学家很感兴趣的扩环产物. 环己酮加氧酶也是有用的生物催化剂, 由于辅酶再生的问题已被工程菌克服了, 所以能像全细胞催化剂那样使用. 对酮包括杂环酮进行Baeyer-Villiger氧化和动态动力学拆分, 放大这种反应作为合成路线是很有前途的.     

Characterization and Crystal Structure of a Robust Cyclohexanone Monooxygenase

Cyclohexanone monooxygenase (CHMO) is a promising biocatalyst for industrial reactions owing to its broad substrate spectrum and excellent regio‐, chemo‐, and enantioselectivity. However, the low stability of many Baeyer–Villiger monooxygenases is an obstacle for their exploitation in industry. Characterization and crystal structure determination of a robust CHMO from Thermocrispum municipale is reported. The enzyme efficiently converts a variety of aliphatic, aromatic, and cyclic ketones, as well as prochiral sulfides. A compact substrate‐binding cavity explains its preference for small rather than bulky substrates. Small‐scale conversions with either purified enzyme or whole cells demonstrated the remarkable properties of this newly discovered CHMO. The exceptional solvent tolerance and thermostability make the enzyme very attractive for biotechnology.     

硫化物的生物氧化成手性亚砜

描述了硫化物不对称生物氧化成手性亚砜, 主要为氯过氧化物酶、环己酮单氧酶催化有机硫化物氧化成具有光学活性的亚砜的两条酶促反应途径.     

Asymmetric ketone reduction by a hyperthermophilic alcohol dehydrogenase. The substrate specificity,enantioselectivity and tolerance of organic solvents

In an effort to search for effective biocatalysts for asymmetric ketone reduction, the substrate specificity and enantioselectivity of an alcohol dehydrogenase from the hyperthermophilic archaeon Pyrococcus furiosus have been evaluated. This hyperthermophilic alcohol dehydrogenase catalyzes the reduction of various ketones including aryl ketones, α- and β-ketoesters. Interestingly, aryl ketones, phenyl-substituted α- and β-ketoesters were reduced to the corresponding chiral alcohols in an enantiomerically pure form, while the substrates lacking phenyl groups were reduced with a moderate enantioselectivity. It thus suggests that a phenyl group next to the carbonyl group could be very helpful for achieving an excellent enantioselectivity, and this could provide valuable guidance for the future application of this useful enzyme through rational substrate engineering. The reaction temperature increased the enzyme activity, but exerted no effect on the enantioselectivity. This alcohol dehydrogenase also showed a high tolerance of organic solvents such as dimethyl sulfoxide, iso-propanol, methyl tert-butyl ether, and hexane, a particularly important and useful feature for the reduction of ketones with a low solubility in aqueous buffers.     

The steroid monooxygenase from Rhodococcus rhodochrous; a versatile biocatalyst

The substrate scope of a steroid monooxygenase (STMO) from Rhodococcus rhodochrous DSM 43269 was investigated for a large range of different ketone substrates. These studies revealed that this enzyme not only oxygenates steroids, but also ketone moieties of a series of other open-chain ketones, such as cyclohexyl methyl ketone, cyclopentyl methyl ketone, and 3-acetylindole. Furthermore, the STMO catalyzed the oxygenation of cyclobutanone derivatives. Comparative biotransformations with recombinant Escherichia coli resting cells harboring the STMO, the cycloalkanone monooxygenase (CAMO) from Cylindrocarpon radicicola or the cyclohexanone monooxygenase (CHMO) from Acinetobacter calcoaceticus revealed that the STMO is enantiodivergent compared to the CHMO-type. Moreover, the STMO resulted in a higher enantiomeric excess of the product lactones compared to the known BVMOs of the same enantiopreference, such as cyclopentanone monooxygenases.     

Towards practical biocatalytic Baeyer-Villiger reactions: applying a thermostable enzyme in the gram-scale synthesis of optically-active lactones in a two-liquid-phase system

Baeyer-Villiger monooxygenases (BVMOs) are extremely promising catalysts useful for enantioselective oxidation reactions of ketones, but organic chemists have not used them widely due to several reasons. These include instability of the enzymes in the case of in vitro and even in vivo systems, reactant/product inhibition, problems with upscaling and the necessity of using specialized equipment. The present study shows that the thermally stable phenylacetone monooxygenase (PAMO) and recently engineered mutants can be used as a practical catalysts for enantioselective Baeyer-Villiger oxidations of several ketones on a preparative scale under in vitro conditions. For this purpose several parameters such as buffer composition, the nature of the solvent system and the co-factor regeneration system were optimized. Overall a fairly versatile and efficient catalytic system for enantioselective laboratory scale BV-oxidations of ketones was developed, which can easily be applied even by those organic chemists who are not well versed in the use of enzymes.     

Copper-catalyzed imination of sulfoxides and sulfides

Sulfoximines and sulfilimines have attracted considerable interest among organic chemists. The Cu(II)-catalyzed imination of sulfoxides and sulfides using various N-fluoro benzenesulfonamides was investigated in this study. The scope of the reaction was demonstrated by using several substituted sulfides and sulfoxides. The flow strategy for the preparation of NH-sulfoximines was also examined. By trapping nitrene intermediates through triphenylphosphine, we found that the reaction was conducted through a metal-nitrene intermediate mechanism.     

What to sacrifice? Fusions of cofactor regenerating enzymes with Baeyer-Villiger monooxygenases and alcohol dehydrogenases for self-sufficient redox biocatalysis

A collection of fusion biocatalysts has been generated that can be used for self-sufficient oxygenations or ketone reductions. These biocatalysts were created by fusing a Baeyer-Villiger monooxygenase (cyclohexanone monooxygenase from Thermocrispum municipale: TmCHMO) or an alcohol dehydrogenase (alcohol dehydrogenase from Lactobacillus brevis: LbADH) with three different cofactor regeneration enzymes (formate dehydrogenase from Burkholderia stabilis: BsFDH; glucose dehydrogenase from Sulfolobus tokodaii: StGDH, and phosphite dehydrogenase from Pseudomonas stutzeri: PsPTDH). Their tolerance against various organic solvents, including a deep eutectic solvent, and their activity and selectivity with a variety of substrates have been studied. Excellent conversions and enantioselectivities were obtained, demonstrating that these engineered fusion enzymes can be used as biocatalysts for the synthesis of (chiral) valuable compounds.     

Titanium Tetrachloride in Organic Synthesis [New synthetic methods (21)]

Titanium tetrachloride can accelerate numerous organic reactions. Valuable syntheses of, e.g., allyl sulfides, amides, enamines, and ketones are based upon transformations of functional groups with TiCl₄. Particular mention should also be made of carbon-carbon linkage with TiCl₄ which permits the synthesis of hydroxy ketones and carbonyl compounds of the Michael adduct type. TiCl₄ reduced in situ is suitable for the reduction of chloroarenes or the linkage of two aldehyde molecules to give an alkene.     

Active site model of cyclohexanone monooxygenase. 3. The case of benzyl sulfides bearing polar groups

The stereopreference of cyclohexanone monooxygenase for a series of benzyl methyl sulfides bearing in the p-position groups with different σ_p values has been rationalized by locating inside the enzyme active site model electron-rich and electron-poor regions.     

Enzymatic kinetic resolution of racemic ketones catalyzed by Baeyer–Villiger monooxygenases

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.     

Asymmetric Claisen Rearrangements on Chiral Vinyl Sulfoxides

Highly diastereoselective Claisen rearrangements of acyclic allyl vinyl ethers bearing a chiral sulfoxide at C‐5 provide γ‐δ‐unsaturated aldehydes or ketones with up to two consecutive asymmetric centers in the molecule whilst preserving a useful vinyl sulfoxide. The reactivity of related vinyl sulfides and sulfones has also been examined in this work.     

Rh(III)-catalyzed annulation of azobenzenes and α-Cl ketones toward 3-acyl-2H-indazoles

Rhodium(III)-catalyzed [4 + 1] cyclization of azobenzenes with α-Cl ketones has been developed. 3-Acyl-2H-indazoles could be easily afforded in up to 97% yields for more than 30 examples. The obtained products are potentially valuable in organic synthesis and drug discovery. This protocol featured with high efficiency, extensive functional group tolerance and mild reaction conditions. The one-step efficient construction of an anti-inflammatory agent confirms the practicability of this procedure.     

Flavins as organocatalysts for environmentally benign molecular transformations

Recent progress in the development of flavin-catalyzed oxidations and related reactions is described with respect to scope, limitation, and reaction mechanism. The 4a-hydroperoxyflavins, which are the most simplified model compounds of flavoenzymes, act as catalytically active species for the oxidation of organic substrates with the help of H(2)O(2) or O(2) as a mild oxidant. This principle behind the simulation of flavoenzymes led to the discovery of a variety of environmentally benign, oxidative transformations of secondary amines to nitrones, tertiary amines to N-oxides, sulfides to sulfoxides, and Baeyer-Villiger oxidations of ketones. Asymmetric oxidation of sulfides can also be performed with several chiral flavin catalysts. One of the fortunate outcomes of this study is the development of an environmentally friendly ("green") method for the "aerobic hydrogenation" of olefins, which is achieved by in situ generation of diimide with the aid of the flavin-catalyzed oxidation of hydrazine under an O(2) atmosphere.     

Exploring the biocatalytic scope of a bacterial flavin-containing monooxygenase

A bacterial flavin-containing monooxygenase (FMO), fused to phosphite dehydrogenase, has been used to explore its biocatalytic potential. The bifunctional biocatalyst could be expressed in high amounts in Escherichia coli and was able to oxidize indole and indole derivatives into a variety of indigo compounds. The monooxygenase also performs the sulfoxidation of a wide range of prochiral sulfides, showing moderate to good enantioselectivities in forming chiral sulfoxides.