Journal of Industrial Microbiology & Biotechnology

, Volume 40, Issue 2, pp 245–255

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

Authors

    • Bioprocess and Scale Up Facility, Defence Research and Development EstablishmentMinistry of Defence (Govt. of India)
  • Anil Kumar Singh
    • Bioprocess and Scale Up Facility, Defence Research and Development EstablishmentMinistry of Defence (Govt. of India)
  • Shweta Saraswat
    • Bioprocess and Scale Up Facility, Defence Research and Development EstablishmentMinistry of Defence (Govt. of India)
  • Saurabh Srivastava
    • Bioprocess and Scale Up Facility, Defence Research and Development EstablishmentMinistry of Defence (Govt. of India)
  • Princi Misra
    • Bioprocess and Scale Up Facility, Defence Research and Development EstablishmentMinistry of Defence (Govt. of India)
  • M. Kameswara Rao
    • Division of Biochemistry, Defence Research and Development EstablishmentMinistry of Defence (Govt. of India)
  • N. Gopalan
    • Vector Management Division, Defence Research and Development EstablishmentMinistry of Defence (Govt. of India)
  • P. V. L. Rao
    • Bioprocess and Scale Up Facility, Defence Research and Development EstablishmentMinistry of Defence (Govt. of India)
Fermentation, Cell Culture and Bioengineering

DOI: 10.1007/s10295-012-1220-z

Cite this article as:
Athmaram, T.N., Singh, A.K., Saraswat, S. et al. J Ind Microbiol Biotechnol (2013) 40: 245. doi:10.1007/s10295-012-1220-z

Abstract

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.

Keywords

Fed batch fermentationPichia pastorisHemagglutininH1N1Swine Origin Influenza A virus

Abbreviations

AOX1

Alcohol oxidase promoter 1

DO

Dissolved oxygen

OD

Optical Density

HA

Hemagglutinin

PBS

Phosphate buffered saline

MALDI TOF/MS

Matrix-assisted laser desorption/ionization Time of Flight/Mass spectrometry

HCCA

α-Cyano-4-hydroxy cinnamic acid

EU

European units

MWCO

Molecular weight cut off

DEAE

Diethylaminoethanol

Copyright information

© Society for Industrial Microbiology and Biotechnology 2012