Production of recombinant human epidermal growth factor in Pichia pastoris

Eissazadeh, Samira; Moeini, Hassan; Dezfouli, Maryam Ghandizadeh; Heidary, Somayyeh; Nelofer, Rubina; Abdullah, Md. Pauzi

doi:10.1016/j.bjm.2016.10.017

Cited by 39 publications

(19 citation statements)

References 27 publications

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“…Residual glycerol in the cultivation was not depleted completely at the beginning of methanol induction, which repressed the production of human cathepsin S. A high cell density was not always consistent with the production of human cathepsin S, especially in the case of excess residual glycerol in the culture medium at the beginning of the induced phase. This phenomenon was in accordance with what has been reported on human epidermal growth factor production [Eissazadeh et al, 2017]. Before the initiation of methanol induction, glycerol should be depleted from the cultivation to provide a fluent expression of human cathepsin S.…”

Section: Discussionsupporting

confidence: 91%

“…3) for the production of human cathepsin S, which is similar to what has been reported for laccase production in the P. pastoris host system [Eissazadeh et al, 2017;Hong et al, 2002]. Figure 3 shows that methanol at a higher concentration than 0.5% (v/v) inhibited P. pastoris growth and repressed the expression of human cathepsin S. The toxicity effect of methanol was very evident at 1%, which critically reduced human cathepsin S expression and P. pastoris growth.…”

Section: Discussionsupporting

confidence: 80%

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High Expression of Human Cathepsin S by Recombinant Pichia pastoris with Cod Skin as an Organic Co-Nitrogen Source

Qin

et al. 2017

Microb Physiol

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Human cathepsin S production by recombinant Pichia pastoris using cod skin as the co-nitrogen source was investigated in this study. The addition of carbon sources of glycerol in the fed-batch phase and of methanol in the induction stage was also investigated. A new approach to the highly expression of human cathepsin S was developed using 90 g/L of cod skin (wet weight). After 24 h of the initial fermentation, 4% glycerol (v/v, glycerol/culture) was added once to enhance the cell density (OD₆₀₀) in the cultivation. Then, adding and maintaining methanol at 0.5% (v/v, methanol/cultivation) after about 48 h of fermentation achieved a high expression of human cathepsin S in a 5-L bioreactor. The results demonstrate that the maximum activity of human cathepsin S in the fermentation supernatant reached 7,152 U/L after 96 h of methanol induction. The methylotrophic yeast P. pastoris grown in the medium containing cod skin (90 g/L) as the co-nitrogen source provided a 21% higher cell density (OD₆₀₀) and 18.3% higher human cathepsin S yield than P. pastoris grown in BMGY medium. For the first time, human cathepsin S was successfully expressed by P. pastoris with cod skin as the co-nitrogen source. The glycerol fed-batch controlling strategy and method of maintaining methanol at a constant concentration of 0.5% (v/v, methanol/cultivation) in the induction stage was efficient for P. pastoris growth and the expression of human cathepsin S.

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Section: Discussionsupporting

confidence: 91%

Section: Discussionsupporting

confidence: 80%

High Expression of Human Cathepsin S by Recombinant Pichia pastoris with Cod Skin as an Organic Co-Nitrogen Source

Qin

et al. 2017

Microb Physiol

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“…The presence of alternative oxidase (AOX) genes makes this methylotrophic yeast able to convert methanol into formaldehyde and hydrogen peroxide, thus utilizing methanol as energy and carbon source . In many previous studies, the classic AOX1 promoter (P AOX1 ), including P AOX1 ‐derived variants, was mostly used for recombinant protein expression in P. pastoris owing to its strong and tightly regulated features .…”

Section: Introductionmentioning

confidence: 99%

“…[1][2][3][4][5][6][7] The presence of alternative oxidase (AOX) genes makes this methylotrophic yeast able to convert methanol into formaldehyde and hydrogen peroxide, thus utilizing methanol as energy and carbon source. [8][9][10] In many previous studies, the classic AOX1 promoter (P AOX1 ), including P AOX1 -derived variants, was mostly used for recombinant protein expression in P. pastoris owing to its strong and tightly regulated features. 2,3 From the industrial perspective, however, the main drawbacks of this type of strain are the high amount of heat production and oxygen requirement as well as the presence of methanol as a toxic and flammable substance in the upstream and sometimes downstream stages.…”

Section: Introductionmentioning

confidence: 99%

Comparative study of μ‐stat methanol feeding control in fed‐batch fermentation of Pichia pastoris producing HBsAg: an open‐loop control versus recurrent artificial neural network‐based feedback control

Beiroti

Hosseini

Aghasadeghi

et al. 2019

J of Chemical Tech & Biotech

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BACKGROUND In recent decades, artificial neural network (ANN) has been shown to be a robust and promising tool in monitoring and controlling bioprocess systems. In a previous study, the authors designed a highly accurate and precise recurrent neural network (RNN) for predicting the biomass amount of recombinant Pichia pastoris Mut+ producing intracellular hepatitis B surface antigen (HBsAg) during fed‐batch methanol fermentation. In the current work, the aim was to compare the production efficiency of HBsAg between conventional predefined μ‐stat methanol feeding control (open‐loop control) – already established in large‐scale production – and methanol feeding based on a μ‐stat feedback control system using RNN. For this purpose, for each methanol feeding strategy, bench‐scale, fed‐batch fermentation processes were carried out twice. RESULTS According to the results, in contrast to the established μ‐stat predefined feeding strategy (open‐loop), the deviation of specific growth rate and biomass in μ‐stat feedback control based on RNN was negligible. Also, in the suggested methanol feeding control strategy, the HBsAg titer, specific productivity and yield between the performed fed‐batch fermentations – unlike the conventional method – were approximately identical, with average values of 110.8 μg mL−1, 1.52 μg g−1 dry biomass and 0.34 μg g−1 MeOH respectively. CONCLUSION Comparing the biomass growth pattern and HBsAg production efficiency with the conventional μ‐stat predefined feeding in open‐loop control, the new proposed feeding control system illustrated significantly high process efficiency. This reliable control system based on ANN can have many applications in the biopharmaceutical industry for the control of process key parameters as well as for enhancing process efficiency. © 2019 Society of Chemical Industry

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“…Recombinant P. pastoris can produce a high level of heterologous proteins in a simple and protein‐free medium, and it also performs post‐translation modification relatively similar to the mammalian cells . The presence of two alcohol oxidase (AOX) genes makes this kind of microorganism able to utilize methanol as carbon and energy source . For recombinant protein expression in P. pastoris , the AOX1 promoter is usually used due to its strong and tightly regulated feature .…”

Section: Introductionmentioning

confidence: 99%

Accurate and cost‐effective prediction of HBsAg titer in industrial scale fermentation process of recombinant Pichia pastoris by using neural network based soft sensor

Hosseini

Javidanbardan

Khatami

2019

Biotech and App Biochem

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In the current work, the attempt was made to apply best-fitted artificial neural network (ANN) architecture and the respective training process for predicting final titer of hepatitis B surface antigen (HBsAg), produced intracellularly by recombinant Pichia pastoris Mut + in the commercial scale. For this purpose, in large-scale fed-batch fermentation, using methanol for HBsAg induction and cell growth, three parameters of average specific growth rate, biomass yield, and dry biomass concentration-in the definite integral form with respect to fermentation time-were selected as input vectors; the final concentration of HBsAg was selected for the ANN output. Used dataset consists of 38 runs from previous batches; feed-forward ANN 3:5:1 with training algorithm of backpropagation based on a Bayesian regularization was trained and tested with a high degree of accuracy. Implementing the verified ANN for predicting the HBsAg titer of the five new fermentation runs, excluded from the dataset, in the full-scale production, the coefficient of regression and root-mean-square error were found to be 0.969299 and 2.716774, respectively. These results suggest that this verified soft sensor could be an excellent alternative for the current relatively expensive and time-intensive analytical techniques such as enzyme-linked immunosorbent assay in the biopharmaceutical industry.

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Production of recombinant human epidermal growth factor in Pichia pastoris

Cited by 39 publications

References 27 publications

High Expression of Human Cathepsin S by Recombinant <b><i>Pichia pastoris</i></b> with Cod Skin as an Organic Co-Nitrogen Source

High Expression of Human Cathepsin S by Recombinant <b><i>Pichia pastoris</i></b> with Cod Skin as an Organic Co-Nitrogen Source

Comparative study of μ‐stat methanol feeding control in fed‐batch fermentation of Pichia pastoris producing HBsAg: an open‐loop control versus recurrent artificial neural network‐based feedback control

Accurate and cost‐effective prediction of HBsAg titer in industrial scale fermentation process of recombinant Pichia pastoris by using neural network based soft sensor

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