Baeyer–Villiger monooxygenases: Recent advances and future challenges

Laboratory of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 4, 9747 AG Groningen, The Netherlands.
Current opinion in chemical biology (Impact Factor: 6.81). 12/2009; 14(2):138-44. DOI: 10.1016/j.cbpa.2009.11.017
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Baeyer-Villiger monooxygenases For many enzyme classes, a wealth of information on, for example, structure and mechanism has been generated in the last few decades. While the first Baeyer-Villiger monooxygenases (BVMOs) were already isolated more than 30 years ago, detailed data on these enzymes were lacking until recently. Over the last years several major scientific breakthroughs, including the elucidation of BVMO crystal structures and the identification of numerous novel BVMOs, have boosted the research on BVMOs. This has led to intensified biocatalytic explorations of novel BVMOs and structure-inspired enzyme redesign. This review provides an overview on the recently gained knowledge on BVMOs and sketches the outlook for future industrial applications of these unique oxidative biocatalysts.

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Available from: Marco W Fraaije, Sep 30, 2015
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    • "c o m / l o c a t e / c e j the biocatalyzer (whole cells) in the aqueous phase [4], a stirred tank partitioning bioreactor using ionic liquids as the dispersed phase [10] and CHMO molecular structure changes to fold the serines susceptible to oxidation inside the enzyme [11]. Despite several experimental studies on the molecular structure of CHMO, its catalytic activity and reaction rates of the intermediate steps in the overall reaction in order to propose a basic bioconversion mechanism [12] [13] [14], there have been few studies on kinetic modeling considering simultaneous substrate, product and oxygen inhibition phenomena. Some pseudo empirical kinetic models have been reported following Michaelis–Menten approach to describe product formation and substrate consumption in monooxygenase kinetics [15] [16] [17]. "
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    ABSTRACT: The aim of the current work was to develop a pseudo intrinsic kinetic model, based on elementary reactions, to describe the behavior of the bioconversion of ketones using Escherichia coli TOP10 pQR239. Since there are no reports of the oxygen inactivation constant in the literature, this study gave new insights to find optimal conditions of a suitable oxygen supply during the bioconversion. In this model the reaction mechanism proposed followed the formalism of Langmuir–Hinshelwood and considered both substrate inhibition and oxygen inactivation by the formation of intermediary complexes. Therefore, approximations of the pseudo equilibrium of reaction rates or steady state intermediary species were not considered, which allowed for identifying the role of each reaction step involved in the bioconversion. This kinetic model adequately described the observations with and without substrate inhibition and/or oxygen inactivation. And the regression and the estimated parameters were statistically significant, making these analyses reliable regarding the kinetic behavior of CHMO. Then, substrate and oxygen affinity and inhibition constants were obtained from the kinetic parameters of the model. It was observed that oxygen and substrate presented similar affinity constant values. The substrate inhibition (KIS) and oxygen inactivation (KIO2) constants were determined to be 9.98 μM and 22.3 μM, respectively, showing that the CHMO enzyme was twice more sensitive to inhibition by an excess of substrate than oxygen.
    Chemical Engineering Journal 03/2014; 240:1–9. DOI:10.1016/j.cej.2013.11.047 · 4.32 Impact Factor
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    • "The target enzyme, CHMO, can be considered as the prototype enzyme for Baeyer–Villiger monooxygenases (BVMOs), an enzyme class that is attracting an ever-increasing interest for being applied in biocatalytic processes [14] [15] [16] [17] [18] [19] [20]. CHMO has been studied for its biocatalytic potential in numerous studies which has revealed that it (1) accepts a wide range of aliphatic substrates, (2) is able to perform chemo-and regioselective oxidations, and (3) yields absolute enantioselectivity for many of the tested conversions [19] [21]. "
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    ABSTRACT: Enzyme stability is an important parameter in biocatalytic applications, and there is a strong need for efficient methods to generate robust enzymes. We investigated whether stabilizing disulfide bonds can be computationally designed based on a model structure. In our approach, unlike in previous disulfide engineering studies, short bonds spanning only a few residues were included. We used cyclohexanone monooxygenase (CHMO), a Baeyer-Villiger monooxygenase (BVMO) from Acinetobacter sp. NCIMB9871 as the target enzyme. This enzyme has been the prototype BVMO for many biocatalytic studies even though it is notoriously labile. After creating a small library of mutant enzymes with introduced cysteine pairs and subsequent screening for improved thermostability, three stabilizing disulfide bonds were identified. The introduced disulfide bonds are all within 12 Å of each other, suggesting this particular region is critical for unfolding. This study shows that stabilizing disulfide bonds do not have to span many residues, as the most stabilizing disulfide bond, L323C-A325C, spans only one residue while it stabilizes the enzyme, as shown by a 6 °C increase in its apparent melting temperature.
    FEBS Open Bio 02/2014; 4. DOI:10.1016/j.fob.2014.01.009 · 1.52 Impact Factor
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    • "Recent advances and future challenges for Baeyer–Villiger monooxygenases (BVMOs) were pointed out by Torres Pazmiño et al. [1]. This review provides an overview on the recently gained knowledge on BVMOs and sketches the outlook for future industrial applications of these unique oxidative biocatalysts. "
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    ABSTRACT: Baeyer–Villiger bioconversion productivity of the cyclic ketone (±)-cis-bicyclo [3.2.0] hept-2-en-6-ona by the biocatalyst Escherichia coli TOP10 pQR239 in a multiphase system can be limited by mass transport. Mass transfer rates through the liquid–liquid interface depend on the volumetric mass transfer coefficient (kA) and the substrate and product partition coefficients. In situ experimental determination of the volumetric mass transfer coefficient in a partitioning bioreactor is complex. In this work, the substrate (kS) and product (kP) global mass transfer coefficients were determined in a modified Lewis cell in three water–ionic liquids systems. The ionic liquids used were butylmethylpyrrolidinium bis(trifluoromethylsulfonyl)imide [MeBuPyrr][BTA], trioctylmethylammonium bis(trifluoromethylsulfonyl)imide [OMA][BTA] and 1-butyl-3-metyl-imidazolium hexafluorophosphate [BMIM][PF6]. The maximum kS and kP values obtained were 4.35 × 10−5 and 1.21 × 10−5 m s−1 for water-[MeBuPyrr][BTA] system; 1.53 × 10−5 and 7.84 × 10−6 m s−1 for water-[OMA][BTA] system, respectively; and kS values up to 1.01 × 10−5 m s−1 were found for the water-[BMIM][PF6] system. The association among the mass transfer coefficients and the physicochemical properties (interfacial tension, viscosity and density) and the thermodynamics (partition coefficients) are analysed and discussed. Finally, the volumetric mass transfer coefficients (kSA and kPA) were calculated using interfacial areas (A) of the dispersed ionic liquid phase estimated from the “Sauter” mean drop diameter (d32) in a one litre stirred tank partitioning bioreactor.
    Chemical Engineering Journal 02/2012; 181(181-182):702-707. DOI:10.1016/j.cej.2011.12.060 · 4.32 Impact Factor
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