Simple high-cell density fed-batch technique for high-level recombinant protein production with Pichia pastoris: Application to intracellular production of Hepatitis B surface antigen.
gurramkonda et al_final.pdf
Open Access publication
Cast your vote
You can rate an item by clicking the amount of stars they wish to award to this item.
When enough users have cast their vote on this item, the average rating will also be shown.
Your vote was cast
Thank you for your feedback
Thank you for your feedback
Nemani, Satish Kumar
MetadataShow full item record
AbstractABSTRACT: BACKGROUND: Hepatitis B is a serious global public health concern. Though a safe and efficacious recombinant vaccine is available, its use in several resource-poor countries is limited by cost. We have investigated the production of Hepatitis B virus surface antigen (HBsAg) using the yeast Pichia pastoris GS115 by inserting the HBsAg gene into the alcohol oxidase 1 locus. RESULTS: Large-scale production was optimized by developing a simple fed-batch process leading to enhanced product titers. Cells were first grown rapidly to high-cell density in a batch process using a simple defined medium with low salt and high glycerol concentrations. Induction of recombinant product synthesis was carried out using rather drastic conditions, namely through the addition of methanol to a final concentration of 6 g L-1. This methanol concentration was kept constant for the remainder of the cultivation through continuous methanol feeding based on the on-line signal of a flame ionization detector employed as methanol analyzer in the off-gas stream. Using this robust feeding protocol, maximum concentrations of ~7 grams HBsAg per liter culture broth were obtained. The amount of soluble HBsAg, competent for assembly into characteristic virus-like particles (VLPs), an attribute critical to its immunogenicity and efficacy as a hepatitis B vaccine, reached 2.3 grams per liter of culture broth. CONCLUSION: In comparison to the highest yields reported so far, our simple cultivation process resulted in an ~7 fold enhancement in total HBsAg production with more than 30% of soluble protein competent for assembly into VLPs. This work opens up the possibility of significantly reducing the cost of vaccine production with implications for expanding hepatitis B vaccination in resource-poor countries.
CitationSimple high-cell density fed-batch technique for high-level recombinant protein production with Pichia pastoris: Application to intracellular production of Hepatitis B surface antigen. 2009, 8:13 Microb. Cell Fact.
AffiliationHelmholtz Centre for Infection Research, Braunschweig, Germany. firstname.lastname@example.org.
JournalMicrobial cell factories
The following license files are associated with this item:
- Application of simple fed-batch technique to high-level secretory production of insulin precursor using Pichia pastoris with subsequent purification and conversion to human insulin.
- Authors: Gurramkonda C, Polez S, Skoko N, Adnan A, Gäbel T, Chugh D, Swaminathan S, Khanna N, Tisminetzky S, Rinas U
- Issue date: 2010 May 12
- Physiological response of Pichia pastoris GS115 to methanol-induced high level production of the Hepatitis B surface antigen: catabolic adaptation, stress responses, and autophagic processes.
- Authors: Vanz AL, Lünsdorf H, Adnan A, Nimtz M, Gurramkonda C, Khanna N, Rinas U
- Issue date: 2012 Aug 8
- Virus-like particle production with yeast: ultrastructural and immunocytochemical insights into Pichia pastoris producing high levels of the hepatitis B surface antigen.
- Authors: Lünsdorf H, Gurramkonda C, Adnan A, Khanna N, Rinas U
- Issue date: 2011 Jun 26
- Expression of hepatitis B surface antigen in the methylotrophic yeast Pichia pastoris using the GAP promoter.
- Authors: Vassileva A, Chugh DA, Swaminathan S, Khanna N
- Issue date: 2001 Jun 1
- Purification of hepatitis B surface antigen virus-like particles from recombinant Pichia pastoris and in vivo analysis of their immunogenic properties.
- Authors: Gurramkonda C, Zahid M, Nemani SK, Adnan A, Gudi SK, Khanna N, Ebensen T, Lünsdorf H, Guzmán CA, Rinas U
- Issue date: 2013 Dec 1