Gurramkonda, Chandrasekhar ; Adnan, Ahmad ; Gabel, Thomas ; Lunsdorf, Heinrich ; Ross, Anton ; Nemani, Satish ; Swaminathan, Sathyamangalam ; Khanna, Navin ; Rinas, Ursula (2009) 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 Microbial Cell Factories, 8 . Article ID 13. ISSN 1475-2859
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Official URL: https://microbialcellfactories.biomedcentral.com/a...
Related URL: http://dx.doi.org/10.1186/1475-2859-8-13
Abstract
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.
Item Type: | Article |
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Source: | Copyright of this article belongs to BioMed Central. |
ID Code: | 109076 |
Deposited On: | 09 Mar 2018 12:12 |
Last Modified: | 09 Mar 2018 12:12 |
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