Process Development for Production of Recombinant Protective Antigen from Bacillus anthracis
* Author to whom correspondence should be addressed

Email: tripathink@gmail.com
Nagesh K. Tripathi*, Ambuj Srivastva and P. V. Lakshmana Rao
Division of Virology, Defence Research and Development Establishment,
Jhansi Road, Gwalior 474 002

Paper received :19.7.06 Revised paper accepted : 2.9.06

Anthrax is caused by Bacillus anthracis, a large gram positive, sporulating, capsulated, non-motile, cylindrical bacterium. Protective antigen (PA) of anthrax toxin is the major component of human anthrax vaccine. Currently available human vaccines against anthrax consist of a cell free filtrate of Bacillus anthracis culture of toxigenic, nonencapsulated strains of Bacillus anthracis. Fermentation method was developed to produce recombinant protective antigen in Bacillus anthracis for use in diagnostic and vaccine studies. For this purpose the recombinant protective antigen protein was produced in 5 L fermentor at 37 0C, pH 7.4 in modified
Luria Bertani media. The dissolved oxygen level was maintained at 30 % of air saturation by control of air flow and agitator speed, and, wherever necessary by enrichment of inlet air with pure oxygen. After fermentation culture was harvested by centrifugation. The culture supernatant from centrifugation was preclarified by 0.22 μm membrane filtration and further concentrated and purified by tangential flow filtration with Biomax-50 membrane. The final yield of purified recombinant protective antigen from fermentations resulted in approximately 485 mg/L of pure biologically active protein. The purity of the recombinant protective antigen was checked by SDS-PAGE analysis and reactivity of this protein was determined by Western blotting and ELISA.

 

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