Background Lactones are important compounds in the field of medicine, material and chemical industry. One of the promising accesses to these flexible scaffolds is NAD(P) + -dependent alcohol dehydrogenases-catalyzed oxidative lactonization of diols, which relies on the construction of an efficient NAD(P) + regeneration system. Results In this study, a novel system combining horse liver alcohol dehydrogenase (HLADH) with the synthetic bridged flavin cofactor was established for biosynthesis of lactones. The reaction conditions of this system were optimized and a variety of lactones including chiral lactones were efficiently obtained from various diols. Compared to the previously reported NAD(P) + -regeneration systems, this system showed better regeneration efficiency and product yield. A two-phase system was further applied to solve the problem of product inhibition, and 80% yield was obtained at the condition of 300 mM substrate. Conclusions This study provides an efficient method to synthesis of lactones from diols under mild conditions. We believe this system will be a promising alternative to promote the synthesis of other valuable compounds. Graphic abstract
Graphical Customizing the enantioselectivity : WT CHMO Acineto leads to enantioselectivity up to 99 % ee ( S ) in the Baeyer-Villiger oxidation of 4-substituted cyclohexanones. By probing the binding pocket of rationally designed variants with different-sized 4-substituted cyclohexanones, single, double and triple mutants were identified as the best mutants providing highly reversed enantioselectivity for these probing molecules. Enzymatic Baeyer-Villiger oxidation provides a promising green route utilizing molecular oxygen as the oxidant to produce chiral lactones. Wild-type (WT) CHMO Acineto leads to enantioselectivity up to 99 % ee ( S ) in the synthesis of substituted ϵ-caprolactones. To reverse the inherent enantiopreference of CHMO Acineto toward an array of cyclohexanones with various chain length, we herein reshaped the binding pocket with a minimal number of mutations by a rational design strategy combining “size-probes” with in silico study, which drastically reduces the screening effort. By probing the binding pocket of variants with different-sized 4-substituted cyclohexanones substrates, single, double and triple mutants were identified as the best mutants providing highly reversed enantioselectivity for these probing molecules, respectively. The successful demonstration of the strategy combining “size-probes” with in silico study in the protein engineering of CHMO Acineto may provide a valuable guidance for facile engineering other BVMOs with customized enantiopreference for the same classified substrates with their substituents on the chiral or prochiral central atom.
危险品化学品经营许可证（带存储）