Preparation, cloning, and high level expression in E. coli of interleukin 2-pseudomonas exotoxin fusion genes.
Using PCR and oligonucleotide-directed mutagenesis, recombinant plasmids which express IL2-PE fusion genes, e.g., IL2-PE40, IL2-PE40 KDEL, IL2-PE66(4Glu), and IL2-PE66(4Glu) KDEL, were constructed and expressed in E. coli at a high level. The fusion proteins were 20-30% of the total soluble bacterial proteins, existing as inclusion bodies in the host cell. In addition, there are single EcoRI, PstI, and SmaI restriction sites at the 5', 3' terminals and the linking region of the fusion genes, respectively, in the constructed expression plasmids so that these vectors could be conveniently converted to other expression vectors containing other cytokines or toxins.
Gao
J
J
First Military Medical University, Guangzhou, China.
Huang
H
H
Wang
X
X
Xu
L
L
Zheng
Z
Z
Liu
X
X
D036002:ADP Ribose Transferases; D001427:Bacterial Toxins / Q000096:biosynthesis* / Q000235:genetics; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005098:Exotoxins / Q000096:biosynthesis* / Q000235:genetics; D005822:Genetic Vectors; D000922:Immunotoxins / Q000235:genetics / Q000378:metabolism*; D007376:Interleukin-2 / Q000096:biosynthesis* / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D010957:Plasmids / Q000235:genetics; D011065:Poly(ADP-ribose) Polymerases / Q000032:analysis; D016133:Polymerase Chain Reaction; D011550:Pseudomonas aeruginosa; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A
D036002:ADP Ribose Transferases; D001427:Bacterial Toxins; D005098:Exotoxins; D000922:Immunotoxins; D007376:Interleukin-2; D011065:Poly(ADP-ribose) Polymerases; D011993:Recombinant Fusion Proteins; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A