Sweet yeast: engineered saccharomyces cerevisiae for commercial brazzein sweetener production

Overview

This project develops a bioprocess for producing brazzein, a naturally sweet-tasting protein, using engineered Saccharomyces cerevisiae yeast as a microbial host. Brazzein offers a sucrose-like taste profile with high potency, excellent solubility, and heat stability, making it an attractive low-calorie sweetener. However, harvesting brazzein from its native Oubli plant source is impractical, and prior attempts at microbial production have not achieved commercial viability. This initiative applies cutting-edge synthetic biology and metabolic engineering to systematically optimize brazzein expression in yeast, delivering a scalable, natural sweetener product referred to as "Sweet Yeast."

Technical specifications

Core technology and approach:

• Modular cloning (MoClo) toolkit for yeast, enabling efficient assembly of promoters, coding sequences, and terminators into expression vectors
• An expanded panel of fourteen characterized secretion signal sequences specifically formatted for S. cerevisiae, with optimal signals already identified for brazzein
• Metabolic engineering strategies including promoter optimization, chaperone and protein disulfide isomerase co-expression, secretory pathway expansion, and proteolytic disruption
• High-dimensional design of experiments facilitated by automation to sample multi-parameter space and achieve global maximum protein expression
• Directed evolution for sweet protein development
• Minimal-purification extraction methods designed to reduce downstream processing costs
• Scale-up to multi-liter fermenters using the SUSFERM facility at University College Cork

Key advantages:

• S. cerevisiae carries "generally recognized as safe" (GRAS) status and is widely used in baking and beverage fermentation
• Well-characterized genome, proteome, and metabolome support predictable engineering
• Suitable for large-scale industrial fermentation

Technology readiness level

The project is at an advanced early-to-mid stage of development. Key enabling methodologies have already been established, including the expanded yeast MoClo toolkit and identification of optimal secretion signals for brazzein, with this work prepared for publication. The team brings extensive experience through a partnership since 2016 with MilisBio Ltd, a start-up focused on novel sweet proteins. The recently established SUSFERM centre at University College Cork provides the fermentation and bioprocessing infrastructure needed for scale-up. Future work will systematically optimize expression constructs, scale production to multi-liter fermenters, and validate extraction and brazzein activity testing toward a commercially viable product.