Substrate oscillations boost recombinant protein release from Escherichia coli

Area of Science:

• Biotechnology
• Bioprocess Engineering
• Microbial Fermentation

Background:

• Intracellular recombinant protein production in prokaryotes requires cell disruption, increasing costs.
• Scalable methods for extracellular protein release are crucial for efficient recovery.
• Modifying culture conditions offers a simpler, scalable alternative to molecular cell design.

Purpose of the Study:

• To quantitatively investigate process technological methods for enhancing extracellular release of periplasmic recombinant proteins.
• To evaluate the impact of oscillatory feeding profiles on recombinant protein release from E. coli.

Main Methods:

• Utilized bioreactor runs to quantitatively analyze protein release under different feeding strategies.
• Implemented a defined oscillatory post-induction feed profile (4 min frequency, triangular oscillation between 45% and 1.3x max feed rate).
• Analyzed primary metabolism, including carbon dioxide yield, to correlate with feeding profiles.

Main Results:

• An oscillatory feeding profile improved alkaline phosphatase release by approximately 60% compared to constant feeding.
• The oscillatory feed profile involved triangular oscillations between 45% and 1.3-fold the maximum feed rate.
• Carbon dioxide yield served as an indicator for identifying preferred feeding profiles.

Conclusions:

• A defined oscillatory feeding profile significantly enhances extracellular recombinant protein release in E. coli.
• This scalable oscillatory feeding strategy has the potential to maximize production rates and reduce costs.
• The approach aligns with Process Analytical Technology (PAT) for science-based process development and control.