Monday, August 2, 2010
Pacific Concourse (Hyatt Regency San Francisco)
Retinoids are represented as an essential compounds in many biological systems. Beta-Carotene is cleaved by beta-carotene monooxygenase (BCMO) to yield two molecules of retinal. Beta-carotene is synthesized from isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) as a building block. IPP and DMAPP can be formed in E. coli via endogenous methyl erythritol phosphate (MEP) pathway and exogenous mevalonate (MVA) pathway. For supplementation of IPP and DMAPP, dxs gene encoding DXP synthase, the rate-limiting step of MEP pathway, was overexpressed, and the optimized MVA pathway was introduced to E. coli. Heterologous crtE, crtB, crtI and crtY genes encoding GGPP synthase, phytoene synthase, phytoene desaturase and lycopene cyclase, were introduced into E. coli, resulting into recombinant strain producing beta-carotene. In order to produce retinal from beta-carotene, BCMO( beta-carotene monooxygenase) gene of marine bacteria was synthesized with codon optimization for E.coli expression and introduced to the b-carotene producing strain. Since retinoids is easily oxidized in aqueous phase, dodecane was overlaid to culture broth to prevent oxidative degradation of retinoids. Under the two-phase culture with equal volume of dodecane and 2% glycerol as carbon source, 135 mg/L retinoids were produced at 72 hour. This work was supported by the Basic Research Program (Grant No. 2009-0084490) of MEST, the EB-NCRC (Grant No. R15-2003-012-02001-0) and BK21 program of Korea