MetGen is excited to announce that it has started the optimization of fermentation conditions to advance the production of targeted EnXylaScope enzymes to pilot scale utilizing ENZINE^® technology platform. The best conditions will be scaled-up for all the four targeted enzymes using pilot fermenter. The scale up work will be used to validate the transferability of lab results to the commercial scale.

In the first phase, MetGen’s work for the optimization of fermentation conditions towards scale-up includes following specific tasks:

• Transformation of MetGen coli production strain with two glucuronidases.
• Flask scale production of the two glucuronidases with MetGen standard operating procedure.
• Development of an internal glucuronidase activity testing method.
• 1L fed-batch production of the two glucuronidases through MetGen standard operating procedure.
• Downstream process development including but not limited to cell lysis, purification through ultrafiltration and endotoxins removal.

In general, MetGen process development stage typically consists of three, progressively larger, fermentation scales where the process development for the specific enzyme is optimized and improvements on the production system are made. The three scales are defined by the total applied volume of the fermentation experiment, 0.2 – 0.5 L (“flask scale”), 1 L (“lab scale”) and 400 L (“pilot scale”). This is to minimize monetary risk and resource expenditure. Each fermentation scale follows the exact same set of process steps to keep them comparable. The fermentation part of the process is a fed-batch cultivation, in which, after a growth phase performed in batch-mode, the production of the target molecule takes place in a feed phase. By tightly controlled feed of the main energy source to the cells, the cell’s metabolism is geared towards enzyme production, rather than further growth. In the fed-batch process used by MetGen, it is essential to develop a proper feed profile: the time course of feeding the limiting substrate, often the energy source, determines to what extent the metabolic machinery of the bacterial cells is used to produce the target molecule. For enzymes it is well known that an optimal process performance is obtained by controlling the cell’s specific growth rate at a certain value. However, this value is product-specific and must be found experimentally. The production strain produces enzyme in the periplasmic space. Therefore, the full fermentation broth is treated with a buffer solution that promotes cell lysis, thus releasing the product in the broth and making it available for harvesting. Lastly, the concentrated enzymatic product is formulated. The formulation has been created for enzyme and microbiological stability. MetGen’s products are supplied as liquid for easy and safe application.