Abstract
An heterologous expression of Vitreoscilla hemoglobin (VHb) for improving cell growth and recombinant protein production has been successfully demonstrated in various hosts, including Pichia pastoris. Lower temperature cultures can enhance target protein production in some studies of P. pastoris. In this study, the strategy of combining heterologous VHb expression and lower temperature cultures in P. pastoris showed that final cell density and viability of VHb+ strain at 23 °C were higher than that at 30 °C. In addition, the effects of VHb expression on recombinant β-galactosidase production and oxygen uptake rate were also higher at 23 °C than at 30 °C. Consequently, lower temperature cultures can enlarge VHb effectiveness on cell performance of P. pastoris. This is because VHb activity obtained at 23 °C cultures was twofold higher than that at 30 °C cultures, due to a different heme production. This strategy makes P. pastoris an excellent expression host particularly suitable for increasing the yields of the low-stability and aggregation-prone recombinant proteins.
Highlights
Vitreoscilla hemoglobin (VHb), which is synthesized by the strict aerobic bacterium Vitreoscilla, aids the survival and growth of this organism in oxygen-poor environments
In our previous study [6], intracellular expression of VHb can improve cell performance and recombinant β-galactosidase production in P. pastoris VHb+ strain cultured at 30 °C, under different aeration conditions
A recombinant P. pastoris strain capable of cytosolic co-expression of VHb and β-galactosidase under the control of AOX1 promoter was constructed in our previous study [6]
Summary
Vitreoscilla hemoglobin (VHb), which is synthesized by the strict aerobic bacterium Vitreoscilla, aids the survival and growth of this organism in oxygen-poor environments. Metabolic engineering studies have demonstrated that intracellular expression of VHb in different hosts, including prokaryotes and eukaryotes, leads to enhanced oxygen availability, increased cell growth, and improved metabolite and recombinant protein production [2,3]. In our previous study [6], intracellular expression of VHb can improve cell performance and recombinant β-galactosidase production in P. pastoris VHb+ strain cultured at 30 °C, under different aeration conditions. Some studies have reported that lower temperature cultures at 20–25 °C in P. pastoris can improve the target protein production, lower the cell lysis, reduce the release of intracellular proteases to culture medium and decrease the proteolytic activity [7,8,9]. The proposed strategy is practical and effective
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