The industry’s need for bacterial secretion solutions

Bacterial systems have many advantages when it comes to recombinant protein production. They are relatively fast and simple to develop, extensively characterized which enables significant optimisation and cultivation costs are much lower than those of other systems. On the other hand, they lack properties that can be crucial depending on the protein of interest, such as the ability to perform post translational modifications. In the case of E. coli, the favourite bacterial production host to this day, another drawback is the lack of an efficient and selective secretion system.

The latter characteristic is of high relevance for the protein production industry when a protein is difficult to purify or requires a particularly low degree of contamination from the host’s DNA or proteins for safety reasons.

A functional secretion system can also be a game-changer for proteins that are prone to misfolding and aggregation. When over-expressing a recombinant gene at very high levels, the corresponding peptide chain might not have time to fold properly during its translation from mRNA to protein. This is especially likely if it requires the help of scaffold proteins which may not be available in sufficient quantities in the production host. In this case, the misfolded proteins tend to aggregate and form inclusion bodies. These inclusion bodies can be purified, broken down and made to undergo a re-folding procedure in order to obtain correctly folded, active proteins. Unfortunately, these downstream processes are time-consuming, costly and often not efficient enough in terms of final protein yields.

By continuously exporting outside the cell the proteins of interest while they are being produced, a secretion system limits disruptive interactions, reduces the risk of accumulation inside the cell and promotes correct folding. At the same time, it simplifies the purification process considerably both in terms of quantity and quality of the final product. Overall, secretion solves production challenges on several levels; this is why this characteristic is of such high value to the industry.

The development of VB Secretion

Because of their importance in recombinant protein production, many academic and industry scientists are working on bacterial secretion systems. It is therefore not a surprise that the solution that Vectron is currently developing, VB Secretion, originated from an academic research group. Exchange of ideas and partnership are crucial for science and innovative technologies.

The basis for Vectron’s VB Secretion technology comes from Professor Kelly Hughes’ team’s ground-breaking idea to use truncated flagella as a selective secretion channel. At the University of Utah, USA, they engineered Salmonella bacteria and successfully implemented a secretion system in this host. The team created a spin-off company from their research, T3S Technologies, and continued to develop the solution in Salmonella.

As a human pathogen and little characterized organism compared to E. coli, Salmonella is rarely – if ever –used for recombinant protein production. T3S Technologies needed to adapt its findings to an organism relevant for the industry. After a short partnership, it became clear that the scientific team at Vectron Biosolutions was highly knowledgeable about gene expression and engineering in E. coli, and a good fit to try and transpose the secretion system from Salmonella to E. coli. Vectron’s Board and executives recognised the potential of this technology and the added value it would bring to the services Vectron already offered, VB Expression and VB Evolution. Vectron Biosolutions acquired T3S Technology in 2021 and immediately started working on the innovative VB Secretion platform.

Achieved proof-of-concept and the work ahead

Adapting the T3S Technology’s system to E.coli is no simple task. It has already involved numerous rounds of strain engineering and trial-and-error optimisation – as is always the case in biology.

To begin with, the team had to make E.coli express the truncated flagella required for the formation of a secretion channel. The second challenge was then to ensure selective secretion of the protein of interest. There, proteins varying in structure, size and folding patterns and representing a wide array of industrially relevant products were chosen as models. Several of these model proteins were successfully secreted and serve as proofs-of-concept for the transfer of the secretion technology from Salmonella to E. coli, such as human growth hormone and the enzyme beta-lactamase.

At this stage in the development program, the evidence shows encouraging yields of >10 mg/L from non-optimized expression strains used in cultivations at the microtiter scale. This would typically correspond to > 100 mg/L from a fed batch process. After a simple purification step that involves separating the medium from the biomass, protein purities of well over 90% are expected.

One of the central tasks remaining to further develop and optimise VB Secretion is to better understand which proteins are more efficiently secreted and why. To increase our knowledge here, we rely on our industrial partners. Each new project helps us optimise our platforms further and customer feedback is vital to ensure that our services match the needs and expectations of the industry. Each market has its own demands, specificities and regulations, it is therefore important for us to instore a good dialogue with our partners from the start.Our objective is to establish a new standard for protein production across the whole industry in the near future, and for that we work hard on developing a technology optimally fitted to answer our clients’ needs. 

Read the original article on Pharma’s Almanac here.