title: Development of a whole-cell biocatalyst co-expressing P450 monooxygenase and glucose dehydrogenase for synthesis of epoxyhexane.
abstract: Oxygenases-based Escherichia coli whole-cell biocatalyst can be applied for catalysis of various commercially interesting reactions that are difficult to achieve with traditional chemical catalysts. However, substrates and products of interest are often toxic to E. coli, causing a disruption of cell membrane. Therefore, organic solvent-tolerant bacteria became an important tool for heterologous expression of such oxygenases. In this study, the organic solvent-tolerant Bacillus subtilis 3C5N was developed as a whole-cell biocatalyst for epoxidation of a toxic terminal alkene, 1-hexene. Comparing to other hosts tested, high level of tolerance towards 1-hexene and a moderately hydrophobic cell surface of B. subtilis 3C5N were suggested to contribute to its higher 1,2-epoxyhexane production. A systematic optimization of reaction conditions such as biocatalyst and substrate concentration resulted in a 3.3-fold increase in the specific rate. Co-expression of glucose dehydrogenase could partly restored NADPH-regenerating ability of the biocatalyst (up to 38 % of the wild type), resulting in approximately 53 % increase in specific rate representing approximately 22-fold increase in product concentration comparing to that obtained prior to an optimization.
authors.affiliation: Graduate Program in Biotechnology, Faculty of Science, Mahidol University, Bangkok, Thailand.
authors.forename: Akasit
authors.initials: A
authors.lastname: Siriphongphaew
authors.forename: Pimpaya
authors.initials: P
authors.lastname: Pisnupong
authors.forename: Jirarut
authors.initials: J
authors.lastname: Wongkongkatep
authors.forename: Pranee
authors.initials: P
authors.lastname: Inprakhon
authors.forename: Alisa S
authors.initials: AS
authors.lastname: Vangnai
authors.forename: Kohsuke
authors.initials: K
authors.lastname: Honda
authors.forename: Hisao
authors.initials: H
authors.lastname: Ohtake
authors.forename: Junichi
authors.initials: J
authors.lastname: Kato
authors.forename: Jun
authors.initials: J
authors.lastname: Ogawa
authors.forename: Sakayu
authors.initials: S
authors.lastname: Shimizu
authors.forename: Vlada B
authors.initials: VB
authors.lastname: Urlacher
authors.forename: Rolf D
authors.initials: RD
authors.lastname: Schmid
authors.forename: Thunyarat
authors.initials: T
authors.lastname: Pongtharangkul
mesh_terms: D001412:Bacillus subtilis / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism*; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004275:DNA, Ribosomal / Q000737:chemistry / Q000235:genetics; D004852:Epoxy Compounds / Q000378:metabolism*; D042843:Glucose 1-Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D006586:Hexanes / Q000378:metabolism*; D008969:Molecular Sequence Data; D009249:NADP / Q000378:metabolism; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D017422:Sequence Analysis, DNA
chemical_list: D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D004852:Epoxy Compounds; D006586:Hexanes; D012336:RNA, Ribosomal, 16S; D009249:NADP; D003577:Cytochrome P-450 Enzyme System; D042843:Glucose 1-Dehydrogenase