Engineering Saccharomyces cerevisiae for the production and secretion of Affibody molecules
Journal article, 2022

BACKGROUND: Affibody molecules are synthetic peptides with a variety of therapeutic and diagnostic applications. To date, Affibody molecules have mainly been produced by the bacterial production host Escherichia coli. There is an interest in exploring alternative production hosts to identify potential improvements in terms of yield, ease of production and purification advantages. In this study, we evaluated the feasibility of Saccharomyces cerevisiae as a production chassis for this group of proteins.
RESULTS: We examined the production of three different Affibody molecules in S. cerevisiae and found that these Affibody molecules were partially degraded. An albumin-binding domain, which may be attached to the Affibody molecules to increase their half-life, was identified to be a substrate for several S. cerevisiae proteases. We tested the removal of three vacuolar proteases, proteinase A, proteinase B and carboxypeptidase Y. Removal of one of these, proteinase A, resulted in intact secretion of one of the targeted Affibody molecules. Removal of either or both of the two additional proteases, carboxypeptidase Y and proteinase B, resulted in intact secretion of the two remaining Affibody molecules. The produced Affibody molecules were verified to bind their target, human HER3, as potently as the corresponding molecules produced in E. coli in an in vitro surface-plasmon resonance binding assay. Finally, we performed a fed-batch fermentation with one of the engineered protease-deficient S. cerevisiae strains and achieved a protein titer of 530 mg Affibody molecule/L.
CONCLUSION: This study shows that engineered S. cerevisiae has a great potential as a production host for recombinant Affibody molecules, reaching a high titer, and for proteins where endotoxin removal could be challenging, the use of S. cerevisiae obviates the need for endotoxin removal from protein produced in E. coli.



Heterologous protein production


Recombinant protein production





Saccharomyces cerevisiae


Veronica Gast

Chalmers, Biology and Biological Engineering, Systems and Synthetic Biology

Anna Sandegren

Affibody AB

Finn Dunås

Affibody AB

Siri Ekblad

Affibody AB

Rezan Güler

Affibody AB

Staffan Thorén

Affibody AB

Marta Tous Mohedano

Mikael Molin

Martin Engqvist

Chalmers, Biology and Biological Engineering, Systems and Synthetic Biology

Verena Siewers

Chalmers, Biology and Biological Engineering, Systems and Synthetic Biology

Microbial Cell Factories

14752859 (eISSN)

Vol. 21 1 36

Subject Categories

Industrial Biotechnology

Biochemistry and Molecular Biology





More information

Latest update