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An unstructured growth model for the recombinant methylotrophic yeast P. pastoris Mut+ expressing the heavy‐chain fragment C of botulinum neurotoxin serotype A (BoNT/A(Hc)), was successfully established in quasi‐steady state fed‐batch fermentations with varying cell densities. The model describes the relationships between specific growth rate and methanol concentration, and the relationships between specific methanol and ammonium consumption rates and specific growth rate under methanol‐limited growth conditions. The maximum specific growth rate (μ) determined from the model was 0.08 h−1 at a methanol concentration of 3.65 g/L, while the actual maximum μ was 0.0709 h−1. The maximum specific methanol consumption rate was 0.0682 g/g WCW/h. From the model, growth can be defined as either methanol‐limited or methanol‐inhibited and is delineated at a methanol concentration of 3.65 g/L. Under inhibited conditions, the observed biomass yield (YX/MeOH) was lower and the maintenance coefficient (mMeOH) was higher than compared to limited methanol conditions. The YX/MeOH decreased and mMeOH increased with increasing methanol concentration under methanol‐inhibited conditions. BoNT/A(Hc) content in cells (α) under inhibited growth was lower than that under limited growth, and decreased with increasing methanol concentration. A maximum α of 1.72 mg/g WCW was achieved at a μ of 0.0267 h−1 and induction time of 12 h. © 2000 John Wiley & Sons, Inc. Biotechnol Bioeng 70: 1–8, 2000.
Biotechnology and Bioengineering – Wiley
Published: Oct 5, 2000
Keywords: Pichia pastoris ; fed‐batch; growth modeling; optimization; fermentation; botulinum neurotoxin
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