[Construction, expression and functional characterization of disulfide-stabilized anti-hepatocarcinoma single chain Fv fused with truncated Pseudomonas exotoxin].
OBJECTIVE
To prepare fusion protein of disulfide-stabilized anti-hepatocarcinoma single chain Fv(dsFv)and truncated Pseudomonas exotoxin(PE38).


METHODS
The gene encoding the fusion protein was cloned into the prokaryotic expression vector pTIG which was then transformed into E.coli origami(DE3). The expressed product was purified through Ni-NTA agarose affinity chromatography column. The cytotoxicity of the purified dsFv-PE38 was detected by MTT colorimetry.


RESULTS
The immunotoxin dsFv PE38 was expressed in E.coli origami(DE3) in a soluble form, which accounted for nearly 21% of the total bacterial soluble proteins. The results of cytotoxic assay showed that the dsFv-PE38 could specifically kill hepatocarcinoma cells, while spared the normal hepatic cells.


CONCLUSIONS
The immunotoxin dsFv-PE38 may have some potential value in the treatment of hepatocarcinoma.
Zhao
Jun
J
Department of Pathology, Fourth Military Medical University,Xi'an 710032, China. cheny_zj@fmmu.edu.cn
Sun
Zhi-wei
ZW
Liu
Yan-fang
YF
Yu
Wei-yuan
WY
D036002:ADP Ribose Transferases / Q000235:genetics* / Q000494:pharmacology; D001427:Bacterial Toxins / Q000235:genetics* / Q000494:pharmacology; D006528:Carcinoma, Hepatocellular / Q000276:immunology* / Q000628:therapy; D004220:Disulfides; D004926:Escherichia coli / Q000235:genetics; D005098:Exotoxins / Q000235:genetics* / Q000494:pharmacology; D006801:Humans; D007128:Immunoglobulin Fragments / Q000235:genetics* / Q000494:pharmacology; D000922:Immunotoxins / Q000494:pharmacology*; D008113:Liver Neoplasms / Q000276:immunology* / Q000628:therapy; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000302:isolation & purification / Q000494:pharmacology; D037521:Virulence Factors / Q000235:genetics* / Q000494:pharmacology; D000097668:Pseudomonas aeruginosa Exotoxin A
D036002:ADP Ribose Transferases; D001427:Bacterial Toxins; D004220:Disulfides; D005098:Exotoxins; D007128:Immunoglobulin Fragments; D000922:Immunotoxins; D011993:Recombinant Fusion Proteins; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A; C055966:immunoglobulin Fv