[Gene cloning, expression of fusion protein TGF alpha-PE 40 and its inhibition activity on cancer cell growth].
Recombinant plasmid pYX382 and pYX3825 were constructed by fusing the cDNA encoding transforming growth factor type alpha (TGF alpha) to Pseudomonas exotoxin gene (PE) in which the cell recognition domain was deleted. The chimeric proteins produced by host E. Coli cells BL21 transformed by plasmid pYX382 and pYX3825 are termed TGF alpha-PE 40 which reacts with antibody against TGF alpha or antibody against PE in immunoblotting to show a 46 kd protein band reflecting the fusion of 56 kD TGF alpha peptide and 40 kD truncated Pseudomonas exotoxin molecule. An additional signal sequence OmpA was inserted into upstream region of TGF alpha cDNA in plasmid pYX3825 resulting in the partly secreting of expression product into medium and periplasm of the cells. TGF alpha-PE 40 was purified from medium by MONO Q ion exchange column and TSK 250 gel filtration column attached to Pharmacia EPLC system. The TGF alpha-PE 40 molecules showed a very strong activities inhibiting the protein synthesis and killing the cancer cells overexpressing EGF receptor on the cell surface.
Xu
Y H
YH
Shanghai Institute of Cell Biology, Chinese Academy of Sciences.
Peng
S F
SF
Jiang
W L
WL
Zhu
Y
Y
Chandhary
V
V
FitzGerald
D
D
Pastan
I
I
D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D066246:ErbB Receptors / Q000235:genetics; D005098:Exotoxins / Q000235:genetics*; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics / Q000473:pathology; D009363:Neoplasm Proteins / Q000096:biosynthesis*; D011549:Pseudomonas / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000494:pharmacology; D016211:Transforming Growth Factor alpha / Q000235:genetics*; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism
D018076:DNA, Complementary; D005098:Exotoxins; D009363:Neoplasm Proteins; D011993:Recombinant Fusion Proteins; D016211:Transforming Growth Factor alpha; D066246:ErbB Receptors