To produce biologicals using living expression systems can be a huge task in the biotech industry. Experts in this field believe that a yield-enhancing expression technology utilizing the yeast called Pichia pastoris is the right way to go. While the first bacteria that comes to your mind is Escherichia coli when you think of biotechnology, remember that this bacteria may not be the right option for producing recombinant proteins.
Its major limitation is that it cannot produce postraductional modifications like glycosylation. Remember that glycosylation is important when it comes to producing therapeutic proteins that are tolerable by your body without experiencing antigenic reactions. This is the reason why other organisms like Pichia pastoris are now being used. This article explains the key things you should know about the Pichia pastoris expression system.
Understanding Pichia pastoris
Pichia pastoris is simply a methylotroph yeast that usually grows with the alcohol methanol as the source of energy. Therefore, you can easily grow Pichia pastoris in cell suspension in a fairly strong methanol solution that would otherwise kill many other microorganisms. The good thing is that this is a system that is quite cheap to set up and even to maintain.
Another good thing about Pichia pastoris expression system is that it’s similar to the well-researched Saccharomyces cerevisiae also known as the baker’s yeast. These two yeast species tend to have similar tolerances and conditions, and culturing, but Pichia pastoris may be easily adopted in labs without any need of specialist equipment.
One of the most key features of the Pichia pastoris system is that they are ideal for scale growth as well as culture utilizing bioreactors. You see, this feature when you combine it with some inexpensive growth medium, it can make Pichia pastoris to be a cost-effective and highly efficient expression system.
Several studies have given good results when Pichia pastoris is used compared to other expression systems. For example, in one study, Pichia pastoris successfully expressed many individual receptors similar to a mammalian cell system and even produced at least twice as many more receptors than Escherichia coli. For most of the receptors, they achieved higher functional expression in Pichia pastoris when compared to the mammalian cell system.
When a receptor expression that was in Pichia pastoris was compared to the insect cells, it showed that there were similar levels of binding affinity and specific activity, though the higher cell densities produced in Pichia pastoris meant that the total yield in the system was at least twice that of the insect cells.
The high yields and low cost alongside the eukaryotic processing means that Pichia pastoris is a great alternative to the mammalian and insect cell systems, especially when it involves the large-scale production of several integral membrane proteins required in structural studies.
Also, Pichia pastoris can secrete a bit of endogenous proteins and there is easy purification of the proteins. All these benefits have shown that Pichia pastoris is a great platform for producing heterologous proteins.
Industrial biotechnology has usually used many bacterial and eukaryotic organisms to produce proteins. Most microbial products have been on the industrial biotechnology market for years now.
With the coming of recombinant DNA technology, it’s now possible to produce various traits for the production of needed compounds into non-natural producers. While there are many microbial production systems, bacteria and yeasts usually share many advantageous features including easy cultivation, single cell growth, and easy genetic manipulation.
In most cases, bacteria have generally been considered to be the most ideal and efficient producers when it comes to heterologous proteins because of several reasons. But standard prokaryotic systems may have some issues when it involves producing human proteins. For instance, bacteria cannot perform some of the most complex post-translational modifications because many proteins need extra processing to be fully active.
Glycosylations that are needed to make sure that there is the right activity and function by influencing the right charge, folding, solubility, in vivo activity, the protein’s serum half live, and many more cannot usually be completely done in bacterial systems. However, fungal expression systems, especially yeast, can grow in fairly defined and cheap media, so you can reduce the production costs. Besides, they are not usually susceptible to contamination, and the yeast cells can be less sensitive because their walls are more resistant to stress while in the production process.
As explained earlier, the Pichia pastoris system is being successfully utilized for the production of multiple recombinant heterologous proteins. Remember that the recombinant protein in the yeast has many benefits compared to other eukaryotic and prokaryotic expression systems. These include rapid growth rate alongside high-density fermentation, a high level of productivity in protein-free medium, elimination of bacteriophage and endotoxin contamination, ease of genetic manipulation, and absence of some known human pathogenicity.
All these benefits make Pichia pastoris an important system for both industrial manufacture and basic laboratory research. You can scale up the fermentation to meet huge demands, and even control parameters that influence protein activity and productivity like carbon source feed rate and pH.
When you compare Pichia pastoris with mammalian cells, you will find out that Pichia pastoris does not need complex culture conditions or growth medium. Instead, it has a eukaryotic protein synthesis system and is genetically simple to manipulate. Also, when it comes to protein purification in Pichia pastoris, it’s usually straightforward. The soluble and secreted proteins may be directly retrieved by clarification of the Pichia pastoris culture medium using centrifugation.
You can concentrate the samples and purify by putting the supernatant to precipitation, ultrafiltration, and adsorption. You can increase the yield of the secreted proteins during fermentation by using a multistage process that can scale up yeast.
This can start from a small starter culture that is in a non-fermenting way, increase biomass through feed-batch fermentation, and enhance the gene of interest. As you can see, the Pichia pastoris expression system is perhaps one of the easiest and cost-effective ways you can get heterologous proteins in industrial technology.