Scalable purification of Bacillus anthracis protective antigen from Escherichia coli.
The anthrax toxin consists of three proteins, protective antigen (PA), lethal factor, and edema factor that are produced by the Gram-positive bacterium, Bacillus anthracis. Current vaccines against anthrax use PA as their primary component. In this study, we developed a scalable process to produce and purify multi-gram quantities of highly pure, recombinant PA (rPA) from Escherichia coli. The rPA protein was produced in a 50-L fermentor and purified to >99% purity using anion-exchange, hydrophobic interaction, and hydroxyapatite chromatography. The final yield of purified rPA from medium cell density fermentations resulted in approximately 2.7 g of rPA per kg of cell paste (approximately 270 mg/L) of highly pure, biologically active rPA protein. The results presented here exhibit the ability to generate multi-gram quantities of rPA from E. coli that may be used for the development of new anthrax vaccines and anthrax therapeutics.
Gwinn
William
W
Human Genome Sciences, Inc., Rockville, MD 20850, USA.
Zhang
Mei
M
Mon
Sandii
S
Sampey
Darryl
D
Zukauskas
David
D
Kassebaum
Corby
C
Zmuda
Jonathan F
JF
Tsai
Amos
A
Laird
Michael W
MW
D000942:Antigens, Bacterial / Q000096:biosynthesis / Q000302:isolation & purification*; D001408:Bacillus anthracis / Q000235:genetics*; D001427:Bacterial Toxins / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D007218:Industrial Microbiology / Q000379:methods; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D012680:Sensitivity and Specificity
D000942:Antigens, Bacterial; D001427:Bacterial Toxins; D011994:Recombinant Proteins