Based on the chemostat results obtained, best culture strategies for both P PDF and P UPP expression systems were also successfully implemented in 15 L fed-batch cultivations where q p and product to biomass yield ( Y P/X *) values were similar than those obtained in chemostat cultivations.
This higher expression was also correlated with a marked upregulation of unfolded protein response (UPR) related genes, likely from an increased protein burden in the endoplasmic reticulum (ER). CALB transcription levels were drastically higher when employing the novel expression systems. Resultsīoth the P PDF and P UPP-based expression systems outperformed similar P GAP-based expression in chemostat cultivations, reaching ninefold higher specific production rates ( q p).
These new alternatives were compared with the classical strong promoter P GAP, using the Candida antarctica lipase B (CalB) as model protein for expression system performance. Most promising conditions were subsequently tested in fed-batch cultivations. Here, a thorough comparative kinetic characterization of two expression systems based on the commercial PDF and UPP promoters (P PDF, P UPP) was first conducted in chemostat cultures. Since promoters play a crucial role in an expression system and the bioprocess efficiency, innovative alternatives are constantly developed and implemented. Pichia pastoris is a powerful and broadly used host for recombinant protein production (RPP), where past bioprocess performance has often been directed with the methanol regulated AOX1 promoter (P AOX1), and the constitutive GAP promoter (P GAP).
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