Large-scale production of yak (Bos grunniens) chymosin A in Pichia pastoris

As a promising biocatalyst, Yarrowia lipolytica lipase 2 (YlLip2) is limited in its industrial applications due to its low thermostability. In this study, a thermostable YlLip2 mutant was overexpressed in Pichia pastoris and its half-life time was over 30 min at 80 °C. To obtain a higher protein secretion level, the gene dosage of the mutated lip2 gene was optimized and the lipase activity was improved by about 89%. Then, the YlLip2 activity of the obtained strain further increased from 482 to 1465 U/mL via optimizing the shaking flask culture conditions. Subsequently, Hac1p and Vitreoscilla hemoglobin (VHb) were coexpressed with the YlLip2 mutant to reduce the endoplasmic reticulum stress and enhance the oxygen uptake efficiency in the recombinant strains, respectively. Furthermore, high-density fermentations were performed in a 3 L bioreactor and the production of the YlLip2 mutant reached 9080 U/mL. The results demonstrated that the expression level of the thermostable YlLip2 mutant was predominantly enhanced via the combination of these strategies in P. pastoris, which forms a consolidated basis for its large-scale production and future industrial applications.

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Cloning, large-scale production and characterization of fusion protein (P-TUFT-ALT-2) of Brugian abundant larval transcript-2 with tuftsin in Pichia pastoris

Preparative Biochemistry & Biotechnology ◽

10.1080/10826068.2018.1514511 ◽

2018 ◽

Vol 48 (9) ◽

pp. 823-833 ◽

Cited By ~ 2

Author(s):

Rajkumar Paul ◽

Selvarajan Karthik ◽

Ponnusamy Vimalraj ◽

Sankaranarayanan Meenakshisundaram ◽

Perumal Kaliraj

Keyword(s):

Pichia Pastoris ◽

Fusion Protein ◽

Large Scale ◽

Scale Production ◽

Large Scale Production

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Assessing virus like particles formation and r-HBsAg aggregation during large scale production of recombinant hepatitis B surface antigen from Pichia pastoris

International Journal of Biological Macromolecules ◽

10.1016/j.ijbiomac.2019.08.019 ◽

2019 ◽

Vol 139 ◽

pp. 697-711

Author(s):

Seyed Nezamedin Hosseini ◽

Taravat Sarvari ◽

Ghazal Bashiri ◽

Maryam Khatami ◽

Seyed Abbas Shojaosadati

Keyword(s):

Pichia Pastoris ◽

Hepatitis B ◽

Surface Antigen ◽

Large Scale ◽

Hepatitis B Surface Antigen ◽

Scale Production ◽

Recombinant Hepatitis ◽

Virus Like Particles ◽

Large Scale Production

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Production of Proteins prM/M and E of Dengue Virus-3 in Pichia pastoris: Simplified Purification and Evaluation of Their Use as Antigens in Serological Diagnosis of Dengue

Fermentation ◽

10.3390/fermentation6030088 ◽

2020 ◽

Vol 6 (3) ◽

pp. 88

Author(s):

Michelle D. O. Teixeira ◽

Roberto S. Dias ◽

John W. O. Prates ◽

Juliana M. C. Monteiro ◽

Mariana F. Xisto ◽

...

Keyword(s):

Pichia Pastoris ◽

Dengue Virus ◽

Ammonium Sulfate ◽

Large Scale ◽

Medical Intervention ◽

Soluble Form ◽

Enzyme Linked Immunosorbent Assay ◽

Scale Production ◽

E Proteins ◽

Large Scale Production

Dengue is a major arbovirus affecting humans today. With the growing number of cases, it is essential to have large-scale production of antigens for the development of diagnostic kits for the rapid detection of patients infected by the virus and consequent proper medical intervention for them. In this work, we express the prM/M and E proteins of dengue virus-3 in yeast Pichia pastoris KM71H. The proteins were produced in soluble form in the supernatant of the culture and were purified by precipitation with ammonium sulfate. The fraction of 80% of ammonium sulfate was used as an antigen in an indirect enzyme-linked immunosorbent assay (ELISA), providing a sensitivity of 82.61% and a specificity of 89.25%. Thus, the methodology proposed here showed promise for obtaining antigens of dengue viruses and creating quick and inexpensive diagnostic tests, which is of great value since large portions of the areas affected by this disease are economically neglected.

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