SBIR Phase I: Platform for Continuous Evolution of Helper Genes to Enable High-Yield Protein Biomanufacturing

The broader/commercial impact of this Small Business Innovation Research (SBIR) Phase I project will be the significant reduction in the cost and complexity of manufacturing high-value biological products. Currently, many potentially life-saving therapeutics, consumer targeted proteins and industrial enzymes are difficult to produce at scale because they are toxic to the production cells or fail to secrete efficiently. Traditional methods to fix these problems are slow and labor-intensive, often taking months to yield only marginal improvements. This project develops a technology that mimics natural evolution at an accelerated pace to rapidly discover genetic solutions that allow cells to manufacture these complex proteins efficiently. By unlocking the production of difficult-to-express proteins, this innovation may lower the cost of biologic drugs, enable novel industrial enzymes for the bio-economy, and strengthen the U.S. position in advanced biomanufacturing. The proposed project aims to develop a scalable, continuous evolution platform capable of engineering yeast strains for high-titer protein secretion. The technical innovation utilizes an orthogonal DNA replication system that allows a specific set of helper genes to mutate at extremely high rates without damaging the host cell's genome. The research objectives for this Phase I project are to validate three proprietary selection systems that couple cell survival to protein secretion and stability. Specifically, the project will evolve yeast strains to tolerate and secrete toxic or unstable benchmark proteins that currently pose manufacturing challenges. The key technical result will be the identification of novel genetic variants that dramatically improve protein yield, and validation of transferability to common production organisms, demonstrating a generalizable workflow for rapid strain engineering that outperforms rational design methods. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.