High-Yield Expression of Gluten-Detoxifying Enzymes in Pichia pastoris for Commercial Gluten-Free Food Production

Project partners: University of Edinburgh & Prozymi Biolabs

Project focus area: Agriculture and food production

This project investigated the production of gluten-detoxifying enzymes in the yeast Pichia pastoris, aiming to support future food and health applications for people affected by gluten-related conditions.

The project partners designed and constructed a series of DNA expression constructs, placing candidate protease enzymes under constitutive, methanol-free yeast promoters and integrating them into the Pichia genome to generate a panel of production strains. Using a standardised small-scale fermentation process, the team assessed how efficiently each strain secreted its target enzyme into the culture medium.

One enzyme, B77, produced a clear protein signal under the tested conditions. Although initial purification attempts using a standard affinity-based method did not yet recover the protein, specific steps for process optimisation were identified for future work.

The remaining enzymes were optimized not only by selecting appropriate induction conditions for protein production, but also through gene optimization, including a gene modifications such the removal of unnecessary cryptic prokaryotic signal sequences.

In parallel, the partners developed a new low-copy plasmid system for co-expressing chaperone proteins KAR2 and PDI, which help fold and secrete challenging proteins. Together, these strains, vectors, and fermentation protocols form a modular toolkit applicable to future enzyme yield improvement, scale-up, and broader industrial applications.

For more information

For more information on this project, contact us, or view all Engineering Biology SPARK Award winners.

This project funding is part of the Engineering Biology Innovation Network, led by Innovate UK Business Connect in collaboration with Innovate UK and UKRI’s Technology Mission Fund. The network’s goal is to progress innovations, create a commercially focused community and foster new consortia to advance innovations towards commercial applications.