title: Optimization of cell density and dilution rate in Pichia pastoris continuous fermentations for production of recombinant proteins.
abstract: This paper provides an approach for optimizing the cell density (Xc) and dilution rate (D) in a chemostat for a Pichia pastoris continuous fermentation for the extracellular production of a recombinant protein, interferon tau (INF-tau). The objective was to maximize the volumetric productivity (Q, mg INF-tau l(-1) h(-1)), which was accomplished using response surface methodology (RSM) to model the response of Q as a function of Xc and D within the ranges 150< or = Xc < or =450 g cells (wet weight) l(-1) and 0.1 microm< or = D< or =0.9 microm (microm=0.0678 h(-1), the maximum specific growth rate obtained from a fed-batch phase controlled with a methanol sensor). The methanol and medium feed rates that resulted in the desired Xc and D were determined based on the mass balance. From the RSM model, the optimal Xc and D were 328.9 g l(-1) and 0.0333 h(-1) for a maximum Q of 2.73 mg l(-1) h(-1). The model of specific production rate (rho, mg INF-tau g(-1) cells h(-1)) was also established and showed the optimal Xc 287.7 g l(-1) and D=0.0361 h(-1) for the maximum rho (predicted to be 8.92 x 10(-3) mg(-1) g(-1) h(-1)). The methanol specific consumption rate (nu, g methanol g(-1) cells h(-1)) was calculated and shown to be independent of the cell density. The relationship between nu and mu (specific growth rate) was the same as that discovered from fed-batch fermentations of the same strain. The approach developed in this study is expected to be applicable to the optimization of continuous fermentations by other microorganisms.
authors.affiliation: Biological Process Development Facility, Department of Chemical Engineering, University of Nebraska-Lincoln, 207 Othmer Hall, Lincoln, NE 68588-0643, USA.
authors.forename: Wenhui
authors.initials: W
authors.lastname: Zhang
authors.forename: Chih-Ping
authors.initials: CP
authors.lastname: Liu
authors.forename: Mehmet
authors.initials: M
authors.lastname: Inan
authors.forename: Michael M
authors.initials: MM
authors.lastname: Meagher
mesh_terms: D018533:Biomass; D019149:Bioreactors; D001709:Biotechnology / Q000379:methods*; D003470:Culture Media / Q000378:metabolism; D005246:Feedback; D005285:Fermentation; D007218:Industrial Microbiology / Q000379:methods; D007370:Interferon Type I / Q000235:genetics / Q000378:metabolism; D008433:Mathematics; D000432:Methanol / Q000378:metabolism; D010843:Pichia / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism*; D011257:Pregnancy Proteins / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D003470:Culture Media; D007370:Interferon Type I; D011257:Pregnancy Proteins; D011994:Recombinant Proteins; C020947:interferon tau; D000432:Methanol