[reaction: see text] An array of random mutants of cyclohexanone monooxygenase (CHMO) from Acinetobacter sp. NCIMB 9871 was screened against a library of structurally diverse ketones for modifications in the substrate acceptance profile and stereopreference of the enzymatic Baeyer-Villiger biooxidation. While the set of mutant biocatalysts was initially evolved for the enantiocomplementary oxidation of 4-hydroxycyclohexanone, improved and/or divergent stereoselectivities were observed for several substrates. In addition, expanded substrate acceptance to facilitate biotransformation of sterically demanding ketones was found.

Whole-cell biotransformations using engineered strains of Escherichia coli expressing cyclopentanone (CPMO) and cyclohexanone monooxygenases (CHMO) of various bacterial origins have been tested for substrate acceptance on tricyclic ketones. Based on the stereopreference of the biocatalytic Baeyer-Villiger oxidation, our recent clustering of this library of enzymes into two distinct groups based on protein sequence was confirmed. Together with short and facile reaction sequences for the production of the substrate ketones, microbial biooxidation enables access to antipodal product lactones as versatile building blocks in natural product and bioactive compound synthesis.The stereoselective Baeyer-Villiger (BV) oxidation to chiral lactone intermediates has received considerable attention in recent years. 1 Both organometal complexes 2 and enzymes (BV monooxygenases -BVMOs) 3 are available as enantioselective catalysts fulfilling the requirements of sustainable and green chemistry strategies.Very recently, we introduced a platform of recombinant strains of Escherichia coli overexpressing BVMOs of various microbial origin. 4,5 This small library of enzymes possesses overlapping substrate specificity for the biooxidation of various ketones and displays enantiocomplementary 6 and regio-divergent biotransformations. 7 The utilization of recombinant whole-cells minimizes potential enzymatic side reactions 8 and simultaneously allows for a facile application of cofactor dependent biocatalysts. 9 Our current research program aims at providing an easy-to-use biocatalytic toolbox of renewable catalytic entities for stereoselective BV oxidations 10 to organic chemists and to demonstrate its potential in natural product synthesis.Enzyme-mediated BV biooxidation of tricyclic bridged ketones of type 1 as prochiral precursors potentially generates four new chiral centers (2) in a single desymmetrization step (Scheme 1). 11Tricyclic bridged ketones 1a,b were prepared according to the literature 12 with minor modifications in a short and efficient sequence: initial Diels-Alder cyclization of the appropriate alkenes with activated diene 3 to 5a,b was followed by reductive removal of the chlorines (6a,b), acid-catalyzed hydrolysis of ketal (7a,b), and subsequent hydrogenation using Pd/C (Scheme 2).The synthetic route to compounds of type 1c,d utilized carbic anhydride 8 to generate diol 9. 13 Subsequent transformations involved activation of the hydroxyls (10) and Kolbe nitrile formation (11). 14 Cyclization to ketone 1c was performed via diacid 12, following a route recently developed by our group for the preparation of fused bicycloketones (Scheme 3). 6c Alkene 1c was easily converted to compound 1d by atmospheric pressure hydrogenation.Only related bicyclo-substrates of type 1a,b were previously oxidized by a cyclohexanone monooxygenase (CHMO) using isolated enzyme. 15 The stereopreference of monooxygenases originating from Acinetobacter (CHMO Acineto ), 16 Arthrobacter 20 and Rhodococcus (CHMO Rhodo1 , CHMO Rhodo2 ) 17 species in...

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