A dynamic fed batch strategy for a Pichia pastoris mixed feed system to increase process understanding

Area of Science:

• Biotechnology
• Microbial Physiology
• Bioprocess Engineering

Background:

• Recombinant Pichia pastoris (P. pastoris) fermentation requires optimized feeding strategies for high productivity.
• Traditional methods like fed-batch or continuous cultivations are time-consuming for parameter optimization.
• Decoupling substrate feeding offers a novel approach for enhanced process understanding.

Purpose of the Study:

• To decouple methanol and glycerol feeding in a mixed substrate fed-batch for P. pastoris.
• To gain deeper process understanding of metabolism and recombinant protein expression.
• To optimize feeding parameters for maximal productivities.

Main Methods:

• Controlled decoupling of methanol and glycerol feeding in a mixed substrate fed-batch.
• Application of a stepwise increased glycerol uptake rate (qGly) control scheme.
• Separate control of specific substrate uptake rates (qs) for parallel characterization.

Main Results:

• A critical specific glycerol uptake rate was identified, beyond which specific productivity declined.
• Dynamic fed-batch approach characterized time-dependent acceleration of protein expression.
• Metabolism and recombinant protein expression were characterized in parallel.

Conclusions:

• The dynamic fed-batch strategy is valuable for P. pastoris process understanding.
• This approach aids in optimizing feeding design for maximal productivities.
• Decoupled substrate feeding enhances recombinant protein production efficiency.