BRIC DOCTORATE PROGRAMME-An upstream platform for the production of high grade heterologous proteins in Pichia pastoris

Porject title- An upstream platfrom for the production of high grade heterologous proteins in Pichia pastoris Background and significance Pichia pastoris is finding increasing use as a host for the expression of a wide variety of recombinant proteins in the laboratory and in industry. It is capable of successfully secreting the product of interest thus easing overall processing. However, the product titres obtained are usually not very high. As a result most processes include high cell density fermentations to allow large quantities of the product to be made. On the other hand, high cell densities are known to result in reduced cell viability, increased cell debris, as well as an increase in the concentration of amorphous and colloidal organic constituents, thus posing a challenge to the purification stage. In addition the high biomass requires a dilution step to accommodate most recovery steps such as centrifugation. Challenges There are several challenges (amongst others) currently facing HCD processing 1. Extension of cell viability 2. Enhancement of protein expression 3. Enhancement / maintenance of product quality Cell viability which impacts on the level of contaminants in broth (e.g.cell debris) and protease action is the result of both cell design and processing conditions namely stress exerted by foreign protein production, length of culture, and potentially, pH levels and adaptability of cell to the inducer. Furthermore, the interrelation between the 'quality' (e.g. robustness and physico-chemical characteristics) of the desired molecule and the process conditions is not yet well established. Objectives a) To investigate the interaction between culture conditions, cell viability distribution and product expression with the view to enhance production ,reduce the burden on downstream processing and achieve optimal cell separation. b)To investigate the impact of the upstream conditions on product quality. Where appropriate the project will draw on existing tool box of technology available in the UCL Advanced Centre for Biochemical Engineering