News & Blogs » Protein News » Why is Yeast Important for Recombinant Protein Expression
Among different bioprocessing technologies for manufacturing functional recombinant therapeutic proteins (e.g., monoclonal antibody drugs, hormones, and vaccines), yeast-based platforms are among the fastest, most flexible, and economically available to researchers from academia to pharma. Aside from Escherichia coli platforms, which provide several advantages over yeast, including a well-known system, easy workflow, and perhaps the most cost-effective, yeast is undoubtedly the most accessible eukaryotic system that may be leveraged for bioprocessing. A definite advantage over bacterial expression systems is the presence of cellular machinery in the yeast ensuring proper protein folding and the addition of post-translational modifications, such as glycosylation (e.g., N-glycosylation and O-glycosylation), acetylation, amidation, hydroxylation, methylation, phosphorylation, pyrrolidone carboxylic acid, sulfation, and ubiquitylation (Gupta and Shukla, 2017, Gomes et al. 2018).
As a recombinant protein expression system, yeast supports the high-yield production of large proteins (i.e., over 50 kDa), and it’s easily genetically modified by various approaches to optimize metabolic, glycosylation, and secretory pathways, ensuring the generation of functional proteins (Demain and Vaishnav, 2009). Additionally, the development of strains harboring expression cassettes with promoters selected through computational and library screen strategies has optimized protein expression in various yeast variants (Madhavan et al. 2021).
|Cost||Production Time||Scale-up Capacity||Propagation||Product Yield||Contamination Risk|
|Bacteria||Low||Low||High||Easy||High||Medium (e.g., endotoxins)|
|Mammalian cells||High||High||Low||Hard||Medium||Very high (e.g., virus, DNA)|
Retrieved and modified from de Sa Magalhaes and Keshavarz-Moore, 2021 (https://creativecommons.org/licenses/by/4.0/)
Saccharomyces cerevisiae and Pichia pastoris, also known asKomagataella phaffii, are the two yeast strains most commonly used for therapeutic protein production. For example, S. cerevisiae has supported the manufacturing of Glucagon-like peptide 2 and IFNα2b, while P. pastoris has been used in the production of hepatitis B surface antigen (HBsAg), human granulocyte-macrophage colony-stimulating factor, insulin, and human serum albumin, among many other therapeutic recombinant proteins (Madhavan et al. 2021). S. cerevisiae’s fast growth and extracellular secretion of expressed recombinant proteins make it an attractive system for bioprocessing (Gupta and Shukla, 2017). Similarly, the production of biological active compounds benefits from P. pastoris’s ability to reach high cell densities in cost-effective culture conditions and express extracellular and intracellular recombinant proteins. Although both S. cerevisiae and P. pastoris may support vaccine development, such as subunit vaccines, P. pastoris’s high-density growth, lower level of protein-glycosylation (i.e., mannose), and higher yields of secreted proteins have made it an ideal candidate for vaccine manufacturing (de Sa Magalhaes and Keshavarz-Moore, 2021, Demain and Vaishnav, 2009).
As a host system for recombinant protein expression, one main advantage of the yeast is being amenable to genetic manipulation. Therefore, to better enable producing proteins with desirable properties, such as the right amount and type of glycosylation, variants of each S. cerevisiae and P. pastoris have been developed. For instance, P. pastoris generally modifies recombinant proteins with shorter high-mannose oligosaccharides, having ~20 mannose residues; nevertheless, hypermannosylation occurs with some proteins (e.g., HIVgp120, and neuraminidase of the A/Victoria/3/75 influenza virus). Therefore, deletion of genes involved in glycosylation may prove an effective strategy, as demonstrated for the PNO1 (Phosphomannosylation of N-linked Oligosaccharides) gene. Deleting PNO1 in P. pastoris helped reduce the extentof glycosylation of human antithrombin, resulting in a product with reduced immunogenicity (Miura et al. 2004, Gomes et al. 2018). Lastly, genetic engineering of secretion factors, such as the alpha-mating factor, a signal peptide within the N-terminus of proteins produced by P. pastoris, has helped increase the secretion of recombinant proteins (e.g., horseradish peroxidase and lipase) (Gomes et al. 2018).
Overall, while yeast strains and variants provide several advantages over bacteria for bioprocessing, expression system selection must be guided by the specific recombinant protein’s requirements. For example, compared to Chinese Hamster Ovarian or CHO cells, yeast cannot introduce some of the more complex glycosylation patterns typically seen in mammalian proteins. While this represents a challenge for therapeutic protein production in this eukaryotic system, different approaches, such as yeast strain selection, gene codon optimization, promoter strength selection, and other genetic engineering strategies, may help achieve functional proteins at high yields (Dalton and Barton, 2014).