simple Pichia pastoris fermentation and downstream processing strategy for making recombinant pandemic Swine Origin Influenza A virus Hemagglutinin protein

• Published in: Journal of industrial microbiology & biotechnology Vol. 40; no. 2; pp. 245 - 255
• Main Authors: Athmaram, T. N, Singh, Anil Kumar, Saraswat, Shweta, Srivastava, Saurabh, Misra, Princi, Kameswara Rao, M, Gopalan, N, Rao, P. V. L
• Format: Journal Article
• Language: English
• Published: Berlin/Heidelberg Springer-Verlag 01-02-2013; Springer; Oxford University Press
• Subjects: Animals; batch fermentation; Biochemistry; Bioinformatics; Biological and medical sciences; Biomass; Biomedical and Life Sciences; Bioreactors; Biotechnology; Cell culture; Cell Culture and Bioengineering; Defense research; Electrophoresis, Polyacrylamide Gel; Endotoxins - analysis; Fermentation; Fundamental and applied biological sciences. Psychology; Genes; Genetic Engineering; Genomes; Glycerol; Hemagglutinin Glycoproteins, Influenza Virus - biosynthesis; Hemagglutinin Glycoproteins, Influenza Virus - genetics; hemagglutinins; Immunization; Influenza; Influenza A virus; Influenza A Virus, H1N1 Subtype - genetics; Influenza A Virus, H1N1 Subtype - metabolism; Inorganic Chemistry; Life Sciences; Manufacturing; Manufacturing industry; Metabolism; Methods. Procedures. Technologies; Microbial engineering. Fermentation and microbial culture technology; Microbiology; Orthomyxoviridae Infections - epidemiology; Orthomyxoviridae Infections - virology; pandemic; Pandemics; Pharmaceutical sciences; Pichia - genetics; Pichia - growth & development; Pichia - metabolism; Pichia pastoris; polyacrylamide gel electrophoresis; Processes; Protein expression; Proteins; R&D; recombinant proteins; Recombinant Proteins - biosynthesis; Recombinant Proteins - genetics; Research & development; sodium dodecyl sulfate; specific growth rate; Studies; swine; Swine - virology; Swine flu; swine influenza; Swine production; Vaccines; Viruses; Western blotting; Yeast; Fed batch fermentation; Hemagglutinin; H1N1; Swine Origin Influenza A virus; Ascomycota; Downstream processing; Orthomyxoviridae; Pichia pastoris; Fermentation; Protein; Pig; Fungi; Virus; Vertebrata; Influenzavirus A; Mammalia; Influenza A virus; Swine; Artiodactyla; Ungulata; Semicontinuous
• ISSN: 1367-5435; 1476-5535
• DOI: 10.1007/s10295-012-1220-z

The present Influenza vaccine manufacturing process has posed a clear impediment to initiation of rapid mass vaccination against spreading pandemic influenza. New vaccine strategies are therefore needed that can accelerate the vaccine production. Pichia offers several advantages for rapid and economical bulk production of recombinant proteins and, hence, can be attractive alternative for producing an effective influenza HA based subunit vaccine. The recombinant Pichia harboring the transgene was subjected to fed-batch fermentation at 10 L scale. A simple fermentation and downstream processing strategy is developed for high-yield secretory expression of the recombinant Hemagglutinin protein of pandemic Swine Origin Influenza A virus using Pichia pastoris via fed-batch fermentation. Expression and purification were optimized and the expressed recombinant Hemagglutinin protein was verified by sodium dodecyl sulfate polyacrylamide gel electrophoresis, Western blot and MALDI-TOF analysis. In this paper, we describe a fed-batch fermentation protocol for the secreted production of Swine Influenza A Hemagglutinin protein in the P. pastoris GS115 strain. We have shown that there is a clear relationship between product yield and specific growth rate. The fed-batch fermentation and downstream processing methods optimized in the present study have immense practical application for high-level production of the recombinant H1N1 HA protein in a cost effective way using P. pastoris.