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Cytotoxic activity of a recombinant GnRH‐PAP fusion toxin on human tumor cell lines
Author(s) -
Schlick Jean-Luc,
Dulieu Philippe,
Desvoyes Bénédicte,
Adami Pascale,
Radom Jean,
Jouvenot Michèle
Publication year - 2000
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(00)01469-1
Subject(s) - immunotoxin , recombinant dna , fusion protein , cytotoxic t cell , ribosome inactivating protein , escherichia coli , receptor , in vitro , biology , cell culture , toxin , microbiology and biotechnology , inclusion bodies , chemistry , cytotoxicity , biochemistry , gene , ribosome , rna , genetics
Pokeweed antiviral protein (PAP), a ribosome‐inactivating protein isolated from the leaves of Phytolacca americana , reveals potent antiviral activity against viruses or cytotoxic action against cells once inside the cytoplasm. Therefore PAP is a good candidate to be used as an immunotoxin. We constructed a bacterial expression plasmid encoding PAP as a fusion protein with gonadotropin‐releasing hormone (GnRH), a neuropeptide with receptor sites on several gynaecologic tumors. The resulting recombinant toxin was produced in Escherichia coli and accumulated in inclusion bodies. After purification under denaturing conditions, renaturated GnRH‐PAP shows an IC 50 of 3 nM on in vitro translation assays and selectively inhibits the growth of the GnRH receptor positive Ishikawa cell line (ID 50 of 15 nM); on the other hand, neither GnRH nor PAP alone had any effect.