Promoter and process engineering for recombinant protein production in Pichia pastoris towards simple, fast and methanol-free cultivation regimes and high product titers
March 4-8, 2018
Protein production in Pichia pastoris often applies methanol-induced gene promoters such as PAOX1 to drive the expression of the target gene. The use of methanol has major drawbacks, so there is a demand for alternative promoters with good induction properties independent of methanol such as the PGTH1 promoter which we reported recently . In order to further increase its potential, we investigated its regulation in more detail by screening of promoter variants harbouring deletions and mutations. Thereby we could identify the main regulatory region and important transcription factor binding sites of PGTH1. We also created a PGTH1 variant, called PG1-3, with greatly enhanced induction properties compared to the wild type promoter.
Model based process engineering could successfully be implemented for PG1-3 to outperform the PAOX1-driven production in a simple feed regime, and to establish a speed fermentation with high titers after only two days total fermentation time.
 Prielhofer, R.; Maurer, M.; Klein, J.; Wenger, J.; Kiziak, C.; Gasser, B.; Mattanovich, D., Induction without methanol: novel regulated promoters enable high-level expression in Pichia pastoris. Microb Cell Fact 2013, 12 (1), 5.
Roland Prielhofer, Michaela Reichinger, Katrien Claes, Christoph Kiziak, Nina Wagner, Brigitte Gasser, and Diethard Mattanovich, "Promoter and process engineering for recombinant protein production in Pichia pastoris towards simple, fast and methanol-free cultivation regimes and high product titers" in "Microbial Engineering", Prof. Eli Keshavarz-Moore , University College London, England Dr. Barry Buckland, BiologicB, USA Eds, ECI Symposium Series, (2018). http://dc.engconfintl.org/microbial/40