Enhancing HSA-GCSFm fusion protein production by Pichia pastoris with an on-line model-based exponential and DO-stat control modes

Evaluation of different glycerol fed-batch strategies in a lab-scale bioreactor for the improved production of a novel engineered β-fructofuranosidase enzyme in Pichia pastoris

“…One of the main findings of that study was the specific growth rate being higher for the narrow DO band (10 − 30% DO) than for the wider DO band (10 − 50% DO) due to the more frequent feeding, thereby corroborating the findings of the current study. These findings are further corroborated by a study by Jia et al, where an online model-based DO-stat control was implemented in the methanol induction stage, to maintain narrower and more stable DO bandwidths (± 20%) (Jia et al 2022 ). The implementation of this control strategy increased the methanol feed, which notably increased cell concentrations as well as heterologous protein production levels (Jia et al 2022 ).…”

“…The methylotrophic yeast Pichia pastoris is recognised as a well-rounded expression system and has successfully been employed as a recombinant host for the expression of a large and increasing number and variety of heterologous proteins, such as Escherichia coli phytase, mouse endostatin, HAS-GCSF m fusion protein and human antigen-binding fragment (Chen et al 2004 ; Garcia-Ortega et al 2013 ; Trollope et al 2015 ; Jia et al 2022 ). This yeast offers many advantages as heterologous protein expression host, which include eukaryotic post-translational modification, growing to high cell-densities on minimal media (maximisation of product yield), high levels of protein expression and secretion as well as ease of genetic manipulation (Karbalaei et al 2020 ; Ergün et al 2021 ; Coetzee et al 2022 ; Pan et al 2022 ).…”

“…However, there is considerable variation in the constant feed rates being employed in recombinant protein production via P. Pastoris , i.e., feed rates from 4 to 161 g/L depending on the protein produced; thus, the effect of constant feed on productivity of a bioprocess merits further investigation (Çalık et al 2015 ). Furthermore, both exponential and constant feed are examples of feed forward control, which means both feeding strategies are based on the cell growth models of the specific organism (Jia et al 2022 ). The specific growth rate for these feeding strategies are not controlled and the growth characteristics of the cultures may change throughout the cultivation process which requires the culture’s feed to be restricted below its optimum specific growth rate (Dietzsch et al 2011 ).…”

Efficient Secretory Expression for Mammalian Hemoglobins in Pichia pastoris

“…Initially, after the first dissolved oxygen rebound, 50% w glycerol was fed at a flow rate of 35 mL per hour for 20 h. After glycerol depletion, t feeding of methanol started when the rebound of dissolved oxygen emerged again at a proximately 48 h to induce the expression of mammalian hemoglobins. The inducti strategy employs the traditional DO-stat control approach, as previously reported [60,6 When the dissolved oxygen (DO) level rose above its upper limit of 20%, the methano feeding medium was initiated to activate the metabolic pathway. The feeding of methan results in a decrease in DO tension to the set value, at which point the feeding is halte Therefore, this method prevented the accumulation of methanol and avoided t The titer of pHB and the SDS-PAGE analysis of the engineered strains.…”

“…Initially, after the first dissolved oxygen rebound, 50% w/v glycerol was fed at a flow rate of 35 mL per hour for 20 h. After glycerol depletion, the feeding of methanol started when the rebound of dissolved oxygen emerged again at approximately 48 h to induce the expression of mammalian hemoglobins. The induction strategy employs the traditional DO-stat control approach, as previously reported [60,61]. When the dissolved oxygen (DO) level rose above its upper limit of 20%, the methanol-feeding medium was initiated to activate the metabolic pathway.…”