Methylotrophic yeastPichia pastoris produced in high-cell-density fermentations with high cell yields as vehicle for recombinant protein production

1989 ◽  
Vol 34 (3) ◽  
pp. 403-404 ◽  
Author(s):  
R. S. Siegel ◽  
R. A. Brierley
Keyword(s):  
Recombinant Protein ◽  
Cell Density ◽  
Recombinant Protein Production ◽  
Protein Production ◽  
High Cell Density ◽  
High Cell

scholarly journals Pichia pastoris: SEL RAGI UNTUK PRODUKSI PROTEIN REKOMBINAN

2018 ◽  
Vol 17 (2) ◽  
Author(s):  
Neng Herawati ◽  
Arizah Kusumawati ◽  
Adi Santoso
Keyword(s):  
Pichia Pastoris ◽  
Recombinant Protein ◽  
Cell Density ◽  
Mammalian Cells ◽  
Recombinant Protein Production ◽  
Protein Production ◽  
High Cell Density ◽  
Expression System ◽  
Scale Up ◽  
High Cell

Pichia pastoris is a group of methylotropic yeast known as a host of expression and protein production which is widely used for biopharmaceutical-based drug production. This yeast can grow fast with a high cell density. Its genetic stability, high cell density, and stress resistance make the development process and scale-up of P. pastoris can increase to a scale of 200,000 liters of culture. In contrast to the expensive and complex development of recombinant protein production in mammalian cells, the development of production in P. pastoris is relatively simple and cheaper. The advantage of P. pastoris as an expression system is that it is able to use methanol as a carbon source by inducing the expression of alcohol oxidase oxidase (AOX) enzyme. Promoter used by this enzyme is also used as a strong promoter for the expression of proteins that we want. Unlike in bacterial and mammalian systems, recombinant protein production in Pichia cells is not contaminated with endotoxins or viruses so it is safer and simplifies the downstream processes in bioproduction. The level of endogenous protein in the low supernatant allows Pichia to cultivate with a high volumetric productivity, therefore the process of protein production becomes very economical. This review provides an overview of several things that must be considered in utilizing P. pastoris as an expression system including the selection of vectors, strains, vector integration mechanisms into the genome, glycosylation processes, and applications in industry.

scholarly journals Proteomic Profiling of Recombinant Escherichia coli in High-Cell-Density Fermentations for Improved Production of an Antibody Fragment Biopharmaceutical

2005 ◽  
Vol 71 (4) ◽  
pp. 1717-1728 ◽  
Author(s):  
Ilana S. Aldor ◽  
Denise C. Krawitz ◽  
William Forrest ◽  
Christina Chen ◽  
Julie C. Nishihara ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Recombinant Protein ◽  
Cell Density ◽  
High Cell Density ◽  
Antibody Fragment ◽  
Culture Conditions ◽  
Test Method ◽  
Differentially Expressed ◽  
Soluble Form ◽  
High Cell

ABSTRACT By using two-dimensional polyacrylamide gel electrophoresis, a proteomic analysis over time was conducted with high-cell-density, industrial, phosphate-limited Escherichia coli fermentations at the 10-liter scale. During production, a recombinant, humanized antibody fragment was secreted and assembled in a soluble form in the periplasm. E. coli protein changes associated with culture conditions were distinguished from protein changes associated with heterologous protein expression. Protein spots were monitored quantitatively and qualitatively. Differentially expressed proteins were quantitatively assessed by using a t-test method with a 1% false discovery rate as a significance criterion. As determined by this criterion, 81 protein spots changed significantly between 14 and 72 h (final time) of the control fermentations (vector only). Qualitative (on-off) comparisons indicated that 20 more protein spots were present only at 14 or 72 h in the control fermentations. These changes reflected physiological responses to the culture conditions. In control and production fermentations at 72 h, 25 protein spots were significantly differentially expressed. In addition, 19 protein spots were present only in control or production fermentations at this time. The quantitative and qualitative changes were attributable to overexpression of recombinant protein. The physiological changes observed during the fermentations included the up-regulation of phosphate starvation proteins and the down-regulation of ribosomal proteins and nucleotide biosynthesis proteins. Synthesis of the stress protein phage shock protein A (PspA) was strongly correlated with synthesis of a recombinant product. This suggested that manipulation of PspA levels might improve the soluble recombinant protein yield in the periplasm for this bioprocess. Indeed, controlled coexpression of PspA during production led to a moderate, but statistically significant, improvement in the yield.

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