Production and purification of a recombinant staphylococcal enterotoxin B vaccine candidate expressed in Escherichia coli

被引:30
作者
Coffman, JD
Zhu, JW
Roach, JM
Bavari, S
Ulrich, RG
Giardina, SL [1 ]
机构
[1] NCI, SAIC Frederick, Biopharmaceut Dev Program, Frederick, MD 21702 USA
[2] USA, Med Res Inst Infect Dis, Ft Detrick, MD 21702 USA
关键词
D O I
10.1006/prep.2001.1556
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
An attenuated, recombinant form of Staphylococcus enterotoxin B (rSEB) was overexpressed in Escherichia coli under transcriptional control of the T7 promoter. The 28-kDa rSEB was partially purified from soluble, intracellular protein by tangential flow filtration and differential ammonium sulfate precipitation. The intermediate product was then further purified using low-pressure liquid chromatography including hydrophobic interaction, cation exchange, and size-exclusion matrices. The final vialed product was >95% pure as determined by Coomassie blue-stained sodium dodecyl sulfate-polyacrylamide gel electrophoresis, high-pressure size-exclusion chromatography, and capillary zonal electrophoresis. The endotoxin level was <0.6 EU/mg. Final estimated yield of purified rSEB was 147 mg/L of starting culture. Purified rSEB was stable, elicited an immune response in mice, and protected mice against a lethal challenge with the native toxin. (C) 2002 Elsevier Science (USA).
引用
收藏
页码:302 / 312
页数:11
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