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Nutrient Regulation by Continuous Feeding for Large-scale Expansion of Mammalian Cells in Spheroids

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Published on: September 25, 2016

Optimizing recombinant mini proinsulin production via response surface method and microbioreactor screening.

Esra Ayan^1,2,3, Ali Özhan Aytekin⁴, Ahmet Kati^2,3

¹Department of Molecular Biology and Genetics, Faculty of Science, Koç University, Istanbul, Türkiye.

|September 8, 2025

Summary

Engineered mini-proinsulin (nMPI) production using a hybrid microscale and statistical modeling approach. Optimized bioprocess achieved high yields, demonstrating a scalable platform for recombinant protein expression in E. coli.

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Area of Science:

Biotechnology
Bioprocess Engineering
Molecular Biology

Background:

Recombinant insulin production requires scalable, high-yield, cost-effective bioprocesses.
Novel mini-proinsulin (nMPI) engineered for enhanced expression and simplified cleavage.
Optimization of nMPI production is crucial for meeting increasing demand.

Purpose of the Study:

To develop and optimize a high-yield, scalable bioprocess for novel mini-proinsulin (nMPI) production.
To apply a hybrid approach combining microscale cultivation and statistical modeling for process optimization.
To validate the predictive accuracy and scalability of the developed optimization system.

Main Methods:

Engineered novel mini-proinsulin (nMPI) with modified C-peptide and residue substitutions.
Utilized BioLector microbioreactor for microscale high-throughput cultivation.
Employed response surface methodology (RSM), including Plackett-Burman Design (PBD) and Central Composite Design (CDD), for medium optimization.
Scaled up the optimized process to a 3-L bioreactor.

Main Results:

Identified glycerol as the most influential medium component for nMPI yield.
Optimized medium formulation (Scenario III) achieved 13.00 g/L productivity in microscale cultivation.
Maintained high performance upon scale-up, reaching 11.5 g/L in a 3-L bioreactor.
Demonstrated balanced carbon and nitrogen sources enhance cell viability and protein expression.

Conclusions:

The hybrid optimization system accurately predicts and scales effectively.
A robust and scalable production platform for nMPI in E. coli was established.
The presented workflow serves as a model for efficient recombinant protein expression systems.
The developed bioprocess is suitable for translation into industrial settings.