Production and purification of Bacillus anthracis protective antigen from Escherichia coli.
Anthrax is caused by the gram-positive, spore-forming bacterium, Bacillus anthracis. The anthrax toxin consists of three proteins, protective antigen (PA), lethal factor, and edema factor. Current vaccines against anthrax use PA as their primary component since it confers protective immunity. In this work, we expressed soluble, recombinant PA in relatively high amounts in the periplasm of E. coli from shake flasks and bioreactors. The PA protein was purified using Q-Sepharose-HP and hydroxyapatite chromatography, and routinely found to be 96-98% pure. Yields of purified PA varied depending on the method of production; however, medium cell density fermentations resulted in approximately 370 mg/L of highly pure biologically active PA protein. These results exhibit the ability to generate gram quantities of PA from E. coli.
Fermentation and Cell Culture Sciences, Human Genome Sciences, Inc., Rockville, MD 20850, USA. mike_laird@hgsi.com
Michael W
MW
Laird
David
D
Zukauskas
Kelly
K
Johnson
Gavin C
GC
Sampey
Henrik
H
Olsen
Andy
A
Garcia
Jeffrey D
JD
Karwoski
Bridget A
BA
Cooksey
Gil H
GH
Choi
Janine
J
Askins
Amos
A
Tsai
Jennifer
J
Pierre
William
W
Gwinn
D000595:Amino Acid Sequence; D000942:Antigens, Bacterial / Q000096:biosynthesis / Q000302:isolation & purification*; D001427:Bacterial Toxins / Q000096:biosynthesis / Q000302:isolation & purification*; D003470:Culture Media; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005285:Fermentation; D008969:Molecular Sequence Data; D019897:Periplasm / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification
D000942:Antigens, Bacterial; D001427:Bacterial Toxins; D003470:Culture Media; D011994:Recombinant Proteins; C030325:anthrax toxin