Development of Sucrose-Fermenting Escherichia Coli K-12 Strains for Low-Cost Overproduction of Recombinant Proteins

Escherichia coli (E. coli) K-12 strains are widely used for overproduction of a variety of recombinant proteins. These strains are incapable of utilizing sucrose as carbon source, therefore, the use of inexpensive raw materials, e.g. molasses, in the fermentation process is not possible. Our goal is to develop a generally applicable model system of E. coli K-12 strains, which expresses recombinant proteins to high levels and is able to grow in media containing sucrose as the main carbon source. The use of sucrose-fermenting E. coli K-12 could substantially reduce the cost of recombinant protein production. In Phase I of the project we will construct and characterize a number of sucrose-fermenting strains, and test production of a model recombinant protein, phosphoenolpyruvate carboxylase. Finally, we will integrate our findings in a genetic-metabolic model, with primary emphasis on defining the optimal genetic arrangement for high-level target protein production. BENEFITS: The primary products resulting from this project will include proprietary genetically engineered E. coli K-12 strains capable of sucrose fermentation and sucrose-rich culture media formulations optimized for these strains. The proposed technology is qenerally applicable for the production_of a variety of recombinant proteins.