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A Lactose-Based Kluyveromyces lactis Cell-Free Protein Synthesis System.

Tejasvi Shivakumar1,2, Akashaditya Das1,3, Maria Victoria Bussoletti Panizo1,2

  • 1Imperial College Centre for Synthetic Biology, Imperial College London, SW7 2AZ London, U.K.

Chem & Bio Engineering
|October 29, 2025
PubMed
Summary

Researchers developed a cell-free protein synthesis (CFPS) platform using Kluyveromyces lactis yeast. This optimized system, using lactose, significantly improved yields for producing therapeutic proteins like Erythropoietin (EPO).

Keywords:
Kluyveromyces lactiscell-free protein synthesisdesign of experimentslactosesynthetic biologyyeast

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Area of Science:

  • Biotechnology
  • Synthetic Biology
  • Metabolic Engineering

Background:

  • Yeasts are known for metabolic capabilities, historically used in food production.
  • Recent advances in metabolic engineering and synthetic biology enable yeast exploitation for pharmaceuticals.

Purpose of the Study:

  • To develop a cell-free protein synthesis (CFPS) platform using Kluyveromyces lactis.
  • To optimize CFPS using lactose as a sustainable carbon and energy source.
  • To demonstrate the platform's utility for producing therapeutic proteins.

Main Methods:

  • Prepared Kluyveromyces lactis extracts for CFPS.
  • Utilized a design-of-experiments (DoE) approach with Latin Hypercube sampling (128 conditions).
  • Tested lactose as a carbon source for biomass and energy in CFPS.

Main Results:

  • Optimized CFPS reaction conditions for Kluyveromyces lactis.
  • Achieved a 4-fold improvement in protein yield compared to baseline.
  • Successfully synthesized deGFP and Erythropoietin (EPO), with EPO yields reaching 54 nM.

Conclusions:

  • Established a yeast-based CFPS platform using Kluyveromyces lactis and lactose.
  • Demonstrated potential for cost-effective biopharmaceutical production.
  • Paved the way for diverse applications, including biosensors and biopharmaceuticals.