The expression of biologically active human p53 in Leishmania cells: a novel eukaryotic system to produce recombinant proteins.
We have investigated the use of Leishmania cells as a novel eukaryotic expression system for the production of recombinant protein. These cells are easy to maintain, requiring no CO2 incubator or shaker, and can be grown in standard tissue culture media. Leishmania cells can be readily transfected with plasmid DNA by electroporation and transformants selected with antibiotic resistance. Recent studies have shown that it is possible to express foreign genes in Leishmania for the purpose of understanding the biology of this protozoan cell. In the present study we report the use of this system as a means of producing a biologically functional human p53 protein. The conformation of the p53 protein is critical for its ability to bind specific DNA sequences. It is demonstrated that Leishmania-synthesized human p53 is phosphorylated and can bind specifically to its enhancer DNA sequence. These data demonstrate that Leishmania may represent a simple eukaryotic expression system for the production of biologically active recombinant proteins.
D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001483:Base Sequence; D003593:Cytoplasm / Q000737:chemistry; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004351:Drug Resistance; D018274:Electroporation; D004742:Enhancer Elements, Genetic; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D005839:Gentamicins / Q000494:pharmacology; D006801:Humans; D007893:Leishmania donovani / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011487:Protein Conformation; D012333:RNA, Messenger / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000032:analysis / Q000096:biosynthesis* / Q000737:chemistry / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000032:analysis / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D000900:Anti-Bacterial Agents; D005839:Gentamicins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D016159:Tumor Suppressor Protein p53; D004247:DNA; C010680:antibiotic G 418