title: Scaling-up Fermentation of Pichia pastoris to demonstration-scale using new methanol-feeding strategy and increased air pressure instead of pure oxygen supplement.
abstract: Scaling-up of high-cell-density fermentation (HCDF) of Pichia pastoris from the lab or pilot scale to the demonstration scale possesses great significance because the latter is the final technological hurdle in the decision to go commercial. However, related investigations have rarely been reported. In this paper, we study the scaling-up processes of a recombinant P. pastoris from the pilot (10 to 100-L) to the demonstration (1,000-L) scales, which can be used to convert 7-β-xylosyl-10-deacetyltaxol into 10-deacetyltaxol by the β-xylosidase for semi-synthesis of Taxol. We demonstrated that a pure oxygen supplement can be omitted from the HCDF if the super atmospheric pressure was increased from 0.05 to 0.10 ± 0.05 MPa, and we developed a new methanol feeding biomass-stat strategy (0.035 mL/g/h) with 1% dissolved oxygen and 100 g/L initial induction biomass (dry cell weight). The scaling-up was reproducible, and the best results were obtained from the 1,000-L scale, featuring a shorter induction time and the highest enzyme activities and productions, respectively. The specific growth and specific production rates were also determined. This study lays a solid foundation for the commercial preparation of 10-deacetyltaxol through the recombinant yeast. It also provides a successful paradigm for scaling-up HCDF of P. pastoris to the demonstration scale.
authors.affiliation: State Key Laboratory of Bioactive Substance and Function of Natural Medicines; Institute of Materia Medica, Chinese Academy of Medical Sciences &Peking Union Medical College, 1 Xian Nong Tan Street, Beijing 100050, P.R. China.
authors.forename: Wan-Cang
authors.initials: WC
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Bioactive Substance and Function of Natural Medicines; Institute of Materia Medica, Chinese Academy of Medical Sciences &Peking Union Medical College, 1 Xian Nong Tan Street, Beijing 100050, P.R. China.
authors.forename: Ting
authors.initials: T
authors.lastname: Gong
authors.affiliation: Key Laboratory of Biosynthesis of Natural Products of National Health and Family Planning Commission, Institute of Materia Medica, Chinese Academy of Medical Sciences &Peking Union Medical College, 1 Xian Nong Tan Street, Beijing 100050, P.R. China.
authors.forename: Qing-Hua
authors.initials: QH
authors.lastname: Wang
authors.affiliation: Key Laboratory of Biosynthesis of Natural Products of National Health and Family Planning Commission, Institute of Materia Medica, Chinese Academy of Medical Sciences &Peking Union Medical College, 1 Xian Nong Tan Street, Beijing 100050, P.R. China.
authors.forename: Xiao
authors.initials: X
authors.lastname: Liang
authors.affiliation: State Key Laboratory of Bioactive Substance and Function of Natural Medicines; Institute of Materia Medica, Chinese Academy of Medical Sciences &Peking Union Medical College, 1 Xian Nong Tan Street, Beijing 100050, P.R. China.
authors.forename: Jing-Jing
authors.initials: JJ
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Bioactive Substance and Function of Natural Medicines; Institute of Materia Medica, Chinese Academy of Medical Sciences &Peking Union Medical College, 1 Xian Nong Tan Street, Beijing 100050, P.R. China.|Key Laboratory of Biosynthesis of Natural Products of National Health and Family Planning Commission, Institute of Materia Medica, Chinese Academy of Medical Sciences &Peking Union Medical College, 1 Xian Nong Tan Street, Beijing 100050, P.R. China.
authors.forename: Ping
authors.initials: P
authors.lastname: Zhu
mesh_terms: D000399:Air Pressure; D061249:Batch Cell Culture Techniques; D018533:Biomass; D005285:Fermentation; D000432:Methanol; D010100:Oxygen / Q000378:metabolism*; D010843:Pichia / Q000378:metabolism*
chemical_list: D010100:Oxygen; D000432:Methanol