Cyclohexanone monooxygenase (E. C. 1.14.13.22) from Acinetobacter sp. NCIB 9871 has been expressed in baker's yeast (Saccharomyces cereVisiae) to create a general reagent for asymmetric Baeyer-Villiger oxidations. This "designer yeast" approach combines the advantages of using purified enzymes (single catalytic species, no overmetabolism, etc.) with the benefits of whole-cell reactions (experimentally simple, no cofactor regeneration necessary, etc.). The yeast reagent was used to systematically examine a series of 2-, 3-, and 4-substituted cyclohexanones (R ) Me, Et, n-Pr, i-Pr, allyl, n-Bu), almost all of which were oxidized to the corresponding -caprolactones in good yields and high enantioselectivities (typically g 95%). Mesomeric 4-substituted cyclohexanones were oxidized to -caprolactones in g 92% ee. The engineered yeast strain also effected kinetic resolutions of 2-substituted cyclohexanones with enantioselectivity values g 200 for substituents larger than methyl. The behavior of 3-substituted cyclohexanones depended upon the size of the substituent. The engineered yeast strain cleanly converted the antipodes of 3-methyl-and 3-ethylcyclohexanone to divergent regioisomers. On the other hand, for cyclohexanones with larger substituents (n-Pr, allyl, n-Bu), both antipodes were oxidized by the enzyme to a single regioisomer. In these cases, the observed enantioselectivities were due to a combination of a modest preference for one enantiomer by the enzyme and an unfavorable conformational preequilibrium required prior to binding of the less-favored antipode, a phenomenon we refer to as substrate-assisted enantioselectivity.

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A wild-type Baeyer-Villiger monooxygenase was engineered to overcome numerous liabilities in order to mediate a commercial oxidation of pyrmetazole to esomeprazole, using air as the terminal oxidant in an almost exclusively aqueous reaction matrix. The developed enzyme and process compares favorably to the incumbent Kagan inspired chemocatalytic oxidation, as esomeprazole was isolated in 87% yield, in >99% purity, with an enantiomeric excess of >99%.

Transition-metal catalysis has emerged as an important means for C-C activation allowing mild and selective transformations. However, the current scope of C-C bonds that can be activated is primarily restricted to either highly strained systems or more polarized C-C bonds. In contrast, catalytic activation of nonpolar and unstrained C-C moieties remains an unmet challenge. Here we report a general approach for catalytic activation of the unstrained C(aryl)-C(aryl) bonds in 2,2’-biphenols. The key is utilizing the phenol moiety as a handle to install phosphinites as a recyclable directing group. Using hydrogen gas as the reductant, mono-phenols are obtained with a low catalyst loading and high functional group tolerance. This approach has also been applied to the synthesis of 2,3,4-trisubstituted phenols. Further mechanistic study suggests that the C-C activation step is mediated by a rhodium(I) mono-hydride species. Finally, a preliminary study on breaking the inert biphenolic moieties in lignin models is illustrated.

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Serratia marcescens has been proved to be a potential strain for industrial 2,3-butanediol production for its high yield, productivity, and other advantages. In this study, the genes slaA, slaB, slaC, and slaR were successfully cloned which were further confirmed to be encoding acetolactate decarboxylase, acetolactate synthase, 2,3-butanediol dehydrogenase, and a LysR-like regulator, respectively. Unlike in Klebsiella sp. or Klebsiella pneumonie and Vibrio sp. or Vibrio cholerae, the gene slaC is separated from other genes. Then it showed that two regulators, SwrR and SlaR, are in charge of this process by exerting effect on the transcription of genes slaA and slaB. By contrast, the expression of gene slaC is unaffected by the two regulators. It means that these two regulators affect the production of 2,3-butanediol by regulating the production of acetoin. Based on these findings, we successfully accelerated the 2,3-butanediol production by inactivation of gene swrR. The obtained results and further investigations should lead to a more suitable fermentation strategy and strain improvement which would be applicable to the industrial production of 2,3-butanediol.

Esomeprazole is the most popular proton pump inhibitor (PPI) for treating gastroesophageal reflux disease. Enzymatic asymmetric sulfoxidation is a green approach to produce chiral sulfoxides. In this report, we focused on optimizing asymmetric sulfoxidation catalyzed by prazole sulfide monooxygenase (AcPSMO). The costly redox cofactor NADPH utilized by AcPSMO was regenerated by formate dehydrogenase with CO 2 as the coproduct, which can be removed easily. During the scale-up process, oxygen supply was found to be the main limiting factor during the early phase of the reaction, while the instability of AcPSMO and the lack of the cofactor NADPH hindered progress during the middle and late phases of the 0.6 L reaction. Finally, by adjusting oxygen mass transfer and increasing the dissolved oxygen, the enzymatic reaction was stepwise amplified to a 120 L scale using a 300 L thermostatic stirred reactor, affording 95.9% conversion and 99.9% enantiomeric excess after 12 h. Extraction and refinement of the product resulted in 0.39 kg of the isolated esomeprazole (sodium salt), with 57.8% overall yield (73.4% before the salt-forming reaction) and 99.1% purity. Thus, a green-by-design system was constructed for the efficient and precise oxidation of omeprazole sulfide into esomeprazole with molecular O 2 as the green cosubstrate and CO 2 and H 2 O as byproducts.

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