title: Optimizing the heterologous production of epothilone D in Myxococcus xanthus.
abstract: The heterologous production of epothilone D in Myxococcus xanthus was improved by 140-fold from an initial titer of 0.16 mg/L with the incorporation of an adsorber resin, the identification of a suitable carbon source, and the implementation of a fed-batch process. To reduce the degradation of epothilone D in the basal medium, XAD-16 (20 g/L) was added to stabilize the secreted product. This greatly facilitated its recovery and enhanced the yield by three-fold. The potential of using oils as a carbon source for cell growth and product formation was also evaluated. From a screen of various oils, methyl oleate was shown to have the greatest impact. At the optimal concentration of 7 mL/L in a batch process, the maximum cell density was increased from 0.4 g dry cell weight (DCW)/L to 2 g DCW/L. Product yield, however, depended on the presence of trace elements in the production medium. With an exogenous supplement of trace metals to the basal medium, the peak epothilone D titer was enhanced eight-fold. This finding demonstrates the significant role of metal ions in cell metabolism and in epothilone biosynthesis. To further increase the product yield, a continuous fed-batch process was used to promote a higher cell density and to maintain an extended production period. The optimized fed-batch cultures consistently yielded a cell density of 7 g DCW/L and an average production titer of 23 mg/L.
authors.affiliation: Department of Process Science, Kosan Biosciences, Inc., 3832 Bay Center Place, Hayward, CA 94545, USA. lau@kosan.com
authors.forename: Janice
authors.initials: J
authors.lastname: Lau
authors.forename: Scott
authors.initials: S
authors.lastname: Frykman
authors.forename: Rika
authors.initials: R
authors.lastname: Regentin
authors.forename: Sally
authors.initials: S
authors.lastname: Ou
authors.forename: Hiroko
authors.initials: H
authors.lastname: Tsuruta
authors.forename: Peter
authors.initials: P
authors.lastname: Licari
mesh_terms: D004355:Drug Stability; D034261:Epothilones; D004852:Epoxy Compounds / Q000378:metabolism*; D005285:Fermentation / Q000502:physiology; D016941:Myxococcus xanthus / Q000254:growth & development / Q000378:metabolism*; D009829:Oleic Acids / Q000378:metabolism*; D012117:Resins, Synthetic; D013379:Substrate Specificity; D013844:Thiazoles / Q000378:metabolism*; D014131:Trace Elements / Q000378:metabolism*
chemical_list: D034261:Epothilones; D004852:Epoxy Compounds; D009829:Oleic Acids; D012117:Resins, Synthetic; D013844:Thiazoles; D014131:Trace Elements; C005576:methyl oleate; C114026:desoxyepothilone B