[Expression and bioactivity of the cloned staphylococcal enterotoxin O].
This study is to clone the gene of staphylococcal enterotoxins O, obtain recombinant protein (rSEO) and investigate its activity on mice lymphocyte. Staphylococcus aureus O gene is cloned into GST gene fusion vector pGEX-4T-1. The resultant plasmid pGEX-4T-SEO was used to transform E. coLi BL21, where the GST-SEO fusion protein was expressed efficiently. Then SEO was purified by Glutathione Sepharose 4B affinity column and digested with thrombin. The bioactivity of SEO was analyzed by MTT assay on mice lymphocyte and tumor cells. The nucleotide sequence was confirmed to code for the protein correctly, and soluble SEO was expressed efficiently in E. coli BL21 with pGEX-4T-SEO. The protein purified by affinity chromatography resulted to be one single band by SDS-PAGE detection. The MTT assay of the purified rSEO demonstrated that its abilities of stimulating T cells and inhibiting the proliferation of K562, K562-ADM and B16 cells were equivalent to that of SEC in vitro. The expression plasmid pGEX-4T-SEO was constructed and the recombinant superantigen was expressed successfully, which may provide a foundation for the further research of the anticancer activity of SEO.
Institute of Pharmacology and Toxicology and Biochemical Pharmaceutics, College of Pharmaceutical Sciences, Zhejiang University, Hangzhou 310058, China.
Ying-Qiu
YQ
Pan
Ding
D
Ding
Hong-Ying
HY
Sun
Shu-Qing
SQ
Chen
D000818:Animals; D049109:Cell Proliferation; D003001:Cloning, Molecular; D004768:Enterotoxins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005260:Female; D005822:Genetic Vectors; D006801:Humans; D020014:K562 Cells; D008214:Lymphocytes / Q000166:cytology; D008297:Male; D008546:Melanoma, Experimental / Q000473:pathology*; D051379:Mice; D008813:Mice, Inbred ICR; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D013154:Spleen / Q000166:cytology; D013211:Staphylococcus aureus / Q000235:genetics; D018089:Superantigens / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*
D004768:Enterotoxins; D011993:Recombinant Fusion Proteins; D018089:Superantigens