(614g) Invited Talk: Metabolic Engineering of Probiotic Yeast for Production of Drugs and Vitamins

The large intestine is the site of many human diseases yet is difficult to access with orally- or intravenously delivered drugs. At the same time, the gut provides a habitat for a numerous and diverse population of microbes (viral, prokaryotic, and eukaryotic) that perform important, health-relevant chemistries using host- and diet-derived carbohydrates as a feedstock. Therefore, there has been a rising interest in genetically engineering these gut microbes to produce therapeutic molecules in the gut, ultimately improving drug delivery and reducing drug costs. Saccharomyces boulardii (Sb) is a widely used yeast probiotic, demonstrating effectiveness against various gastrointestinal disorders. As a yeast, Sb exhibits high rates of protein secretion, is tolerant to low pH, can be freeze-dried, and low (if any) rates of horizontal gene transfer. Sb is therefore a promising chassis for development of personalized engineered probiotic medicines. In this talk, I will discuss our lab’s recent progress in tuning four key parameters of Sb’s efficacy as a drug delivery vehicle. First, I will present the development of a constitutive Sb promoter library, and how we applied this library to synthesize β-carotene in the mouse gut. Then, I will introduce how sensing of extracellular metabolites may be achieved through repair of Sb’s defective mating pathway and the expression of human adenosine and melatonin G protein-coupled receptors. I will next detail our efforts to enhance Sb’s in vivo residence time through both rational and screening-based approaches. Finally, I will discuss some new work from our lab focusing on the engineering and optimization of protein secretion titers in Sb, culminating in a quadruple knockout strain with a 10-fold increase to recombinant peptide production. Taken together, this work establishes Sb as a genetically tractable commensal fungus and demonstrates the tunable delivery of small-molecule and protein therapeutics during colonization.