The design and optimization of biosynthetic pathways for industrially relevant, non-model organisms is challenging due to transformation idiosyncrasies, reduced numbers of validated genetic parts, and a lack of high-throughput workflows. Here we describe a platform for in vitro prototyping and rapid optimization of biosynthetic enzymes (iPROBE) to accelerate this process. In iPROBE, cell lysates are enriched with biosynthetic enzymes by cell-free protein synthesis and then metabolic pathways are assembled in a mix-and-match fashion to assess pathway performance. In this presentation, I will describe the use of cell-free systems coupled to machine learning for optimizing the biosynthesis of 3-hydroxybutyrate, acetone, and products from reverse beta oxidation. We then show how iPROBE selected pathways can be used in <i>Clostridium autoethanogenum</i> to produce bioproducts solely from carbon waste gas. We expect iPROBE coupled to machine learning to accelerate design–build–test cycles for industrial biotechnology.

Cell-free prototyping and rapid optimization of biosynthetic enzymes for cellular design

ACS Fall 2021

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.