Monday, August 13, 2012
Columbia Hall, Terrace Level (Washington Hilton)
There is now a high demand for proteins in industry, especially glycosylated therapeutic proteins and vaccines produced by mammalian cells. As bacterial cells cannot glycosylate proteins and mammalian cells grow slowly, the industry may need to turn to a new option. Tetrahymena thermophila is a ciliated protozoan that grows within days in inexpensive medium and produces glycosylated proteins. Thus, it is a potential candidate for industrial-scale protein production. A novel line of T. thermophila, the SP1-ENT, has been transfected with a plasmid containing the gene encoding bovine enterokinase (ENT) under the control of a starvation-induced promoter. During induction, the medium is replaced with a nutrient-lacking Tris buffer, starving the cells and activating the promoter to induce ENT production. ENT is similar in size and complexity to the malaria vaccine that can be produced by SP1 cells in the future. Throughout growth and ENT protein production, the cells will be monitored in real-time via a novel shake flask platform where each shake flask is equipped with both pH and dissolved oxygen sensors.
The filling ratio, the ratio of culture volume to shake flask volume, directly influences oxygen mass transfer and shear. This study focuses on determining the effect of filling ratio on SP1 ENT production. By assaying the specific activity of ENT produced by cells grown in different filling ratios, we aim to determine the best filling ratio for ENT production. Our initial findings show that filling ratios between 20% and 48% permit the fastest and greatest ENT production.