Dye-Ligand Affinity Chromatography를 이용한 Staphylococcus aureus의 장독소 B 분리 = Purification of Staphylococcal Enterotoxin B by Dye-Ligand Affinity Chromatography

다국어 초록 (Multilingual Abstract)

Staphylococcal enterotoxins, which cause staphylococcal food poisoning, have been purified by several methods. These procedures have usually included various combinations of ion-exchange chromatogram and gel filtration; such procedures can be time-consuming and may also result in low recoveries of enterotoxins. Although high recoveries of staphylococcal enterotoxins have been reported with chromatofocusing, this technique is expensive and cannot easily be adapted for large-scale purification. In this study we applied a simple, single-step procedure for the purification of Staphylococcal enterotoxin B (SEB) from culture supernatant fluids, namely, dye ligand affinity chromatography in which dye is coupled to an agarose support matrix. Dye ligand affinity chromatography has been used to purify a wide range of enzymes and some blood proteins. We used ten kinds of dye ligand affinity columns. The results showed that SEB had the binding ability to Red A, Reactive Green 19, Reactive Green 5, Reactive Brown 10 and Cibacron Blue 3GA-Agarose. We had the highest resolution by using the Reactive Green 19 dye affinity column. The purified SEB produced a single band when subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE) and western blot using anti-SEB. We could also confirm that the purified SEB produced superantigenic effect. Therefore, it could be concluded that SEB was produced purely in single step process by dye-ligand affinity chromatography. Such a method will form the basis of a high-yielding, rapid means of purifying the staphylococcal enterotoxins that can easily be adapted to large-scale production.