This technology is a chemically defined medium for culturing Pichia pastoris that has potential applications in commercial enzyme production and biotherapeutic manufacturing.
Recombinant protein production has revolutionized biotechnology and medicine by facilitating large-scale manufacturing of enzymes and biotherapeutics. Pichia pastoris (Pichia), a species of yeast, has recently gained popularity as a protein expression system due to its rapid growth, efficient protein secretion, maintenance of protein structure, and glycosylation capabilities. However, the media used to grow Pichia remains sub-optimal for commercial protein production. Complete growth media supports rapid Pichia growth, but has components with undefined biologically-derived components such as yeast extract. These undefined components can lead to significant batch-to-batch variation and requires complicated regulatory documentation. Chemically defined Pichia growth media have been also been developed, however, all current formulations significantly decrease both the Pichia growth rate and per-cell productivity. There is therefore a significant need for improved Pichia cultivation media.
These inventors used a combination of rational design and unbiased transcriptomics to develop a chemically defined growth media for Pichia pastoris. Complete medium increases growth rate by supplying organisms with important metabolic intermediates so that the organisms do not need to use energy to synthesize their own. The inventors therefore determined the rate-limiting metabolic intermediates for Pichia growth and designed growth media that provides these nutrients. This medium, called rich defined medium (RDM), contains defined concentrations of glutamine, arginine, asparagine, vitamins, and fatty acids. Growth in RDM media increases the growth rate of Pichia to nearly the same rate as complete medium. Strikingly, RDM also led to substantial improvements in protein expression, and Pichia grown in RDM were shown to produce more heterologous protein per cell than those grown in complete media.
- Defined medium for Pichia pastoris cultivation
- Supports rapid growth rates
- High levels of per-cell protein production