Yeast
Protein production in Pichia pastoris. Yeast is the simplest eukaryotic cell for protein production where the strains Saccharomyces cerevisiae and Pichia pastoris are commonly used. A large variety of eukaryotic proteins are produced to high levels in their functional form in the yeast cell. The main strength with yeast is that it is a simple unicellular eukaryotic cell that is easy, fast, safe and non-expensive to grow with most post-translational modifications possible. The main limitation is the glycosylation pattern which differs from the one in human cells, and this could be of importance for certain target proteins.
P. pastoris is a methylotrophic yeast that is highly suitable for recombinant protein production, mainly due to the expression from the very strong and inducible Alcohol Oxidase Promoter combined with the capacity to grow to high cell densities. This host therefore gives the possibility to produce large amounts of cells in a relatively small volume combined with a high production level per cell; an ideal starting point for subsequent protein purification. Importantly, this also increases reproducibility since one growth is enough for several rounds of purification.
Main advantages with P. pastoris is the ease of secretion as well as efficient production of integral membrane proteins. In conclusion, this yeast is highly suitable for production of eukaryotic proteins where secretion and labelling are established procedures. The Yeast Protein Production (YPP) platform of PPS builds upon protein production in P. pastoris that has been practiced in the Biochemistry research division at the University of Gothenburg for more than 20 years, mainly driven by the research of Kristina Hedfalk. Over the years, a wide variety of eukaryotic proteins have been produced for structural biology, biomaterial as well as medical applications and yields over 100 mg/L have been achieved for integral membrane proteins. A typical workflow for protein production in P. pastoris is illustrated in Figure 1.
What PPS offers:
Being part of PPS, yeast protein production ideally complements the other host systems provided, giving a complete portfolio for the infrastructure and the system will be developed for a wider use of the P. pastoris production system for various applications. Since P. pastoris has shown to be well-suited for production of eukaryotic membrane proteins, the service includes verification of membrane localization and detergent screens for solubilization and purification. Furthermore, all instrumentation for the whole process from DNA design to growth and purification is available using reliable protocols for small-scale screening for high yielding clones, up-scaled growth in fermenters as well as subsequent protein purification and characterizations.
Figure 1.
Schematic representation of the workflow of protein production for P. pastoris.
