title: Scalable purification of Bacillus anthracis protective antigen from Escherichia coli.
abstract: 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.
authors.affiliation: Human Genome Sciences, Inc., Rockville, MD 20850, USA.
authors.forename: William
authors.initials: W
authors.lastname: Gwinn
authors.forename: Mei
authors.initials: M
authors.lastname: Zhang
authors.forename: Sandii
authors.initials: S
authors.lastname: Mon
authors.forename: Darryl
authors.initials: D
authors.lastname: Sampey
authors.forename: David
authors.initials: D
authors.lastname: Zukauskas
authors.forename: Corby
authors.initials: C
authors.lastname: Kassebaum
authors.forename: Jonathan F
authors.initials: JF
authors.lastname: Zmuda
authors.forename: Amos
authors.initials: A
authors.lastname: Tsai
authors.forename: Michael W
authors.initials: MW
authors.lastname: Laird
mesh_terms: 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
chemical_list: D000942:Antigens, Bacterial; D001427:Bacterial Toxins; D011994:Recombinant Proteins