Enhanced heterologous protein production in Pichia pastoris under increased air pressure

Abstract

Pichia pastoris is a widely used host for the production of heterologous proteins. In this case, high cell densities are needed and oxygen is a major limiting factor. The increased air pressure could be used to improve the oxygen solubility in the medium and to reach the high oxygen demand of methanol metabolism. In this study, two P. pastoris strains producing two different recombinant proteins, one intracellular (β-galactosidase) and other extracellular (frutalin), were used to investigate the effect of increased air pressure on yeast growth in glycerol and heterologous protein production, using the methanol AOX1-inducible system. Experiments were carried out in a stainless steel bioreactor under total air pressure of 1 bar and 5 bar. The use of an air pressure raise of up to 5 bar proved to be applicable for P. pastoris cultivation. Moreover, no effects on the kinetic growth parameters and methanol utilization (Mut) phenotype of strains were found, while an increase in recombinant β-galactosidase-specific activity (ninefold) and recombinant frutalin production was observed. Furthermore, the air pressure raise led to a reduction in the secreted protease specific activity. This work shows for the first time that the application of an air pressure of 5 bar may be used as a strategy to decrease protease secretion and improve recombinant protein production in P. pastoris.

Maximizing recombinant human serum albumin production in a MutsPichia pastoris strain

“…Compared to other yeast expression systems, Pichia expression system has several advantages for recombinant protein production such as high cell density, genetic stability, low endogenous protein secretion, posttranslational modification (glycosylation and disulfide bond formation), and high secretory capacity . Thus, P. pastoris has been widely used for recombinant protein production along with bioprocess optimization that is specific to a protein of interest …”

Recombinant O-mannosylated protein production (PstS-1) from Mycobacterium tuberculosis in Pichia pastoris (Komagataella phaffii) as a tool to study tuberculosis infection

“…After the induction in the BSFs, relatively low-levels of extracellular host proteins were detected (Fig. 2a, time 0 post-induction), which is one advantage of producing recombinant proteins in P. pastoris [66, 90].

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Evaluation of recombinant MGL_1304 produced by Pichia pastoris for clinical application to sweat allergy

“…The recombinant protein is produced intracellularly or extracellularly, with many eukaryotic posttranslational modifications such as glycosylation, disulfide bond formation and proteolytic processing. 12,13 In comparison with other protein expression systems such as mammalian cells or insect cells, it is superior in terms of costs and yield of proteins.…”

“…pastoris is a methylotrophic yeast which widely used for heterologous protein expression. 12,13 The AOX1 gene of P. pastoris metabolizes methanol in the medium as its sole source of carbon. P. pastoris is easy to culture, and the recombinant protein is readily purified in large quantities at a low cost, because almost no other exogenous proteins are contained in culture medium for P. pastoris.…”