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.
Laird
Michael W
MW
Fermentation and Cell Culture Sciences, Human Genome Sciences, Inc., Rockville, MD 20850, USA. mike_laird@hgsi.com
Zukauskas
David
D
Johnson
Kelly
K
Sampey
Gavin C
GC
Olsen
Henrik
H
Garcia
Andy
A
Karwoski
Jeffrey D
JD
Cooksey
Bridget A
BA
Choi
Gil H
GH
Askins
Janine
J
Tsai
Amos
A
Pierre
Jennifer
J
Gwinn
William
W
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