The expression of saporin, a ribosome-inactivating protein from the plant Saponaria officinalis, in Escherichia coli

Isabel Barthelemy, Darlene Martineau, Michael Ong, Risë Matsunami, Nicholas Ling, Luca Benatti, Ugo Cavallaro, Marco Soria, Douglas A. Lappi

Research output: Contribution to journalArticlepeer-review

Abstract

We have isolated and sequenced genomic clones from the DNA of Saponaria officinalis using a cDNA probe that encodes proteins with high homology to saporin-6, one of the most potent of the ribosome-inactivating proteins that is currently used for the construction of immunotoxins and mitotoxins. Sequence differences in the clones suggest a multigene family of proteins. These data agree with observations of several different proteins with ribosome-inactivating protein activity and similar structure. Two of the genomic clones encode proteins that have identical sequences to two of the four isoforms of saporin-6. We have inserted the DNA from one genomic clone into an Escherichia coli expression system that encodes a signal sequence for export to the bacterial periplasmic space. Exportation is observed and the isolated gene product has ribosome-inactivation activity similar to the native protein. Sequence analysis shows differential processing of the remaining plant signal sequence. The majority of the expressed protein remains intracellular and this material also shows high specific activity and differential processing. Saporin as an immunotoxin in clinical trial and as a mitotoxin in experimental models has been extremely efficacious. These data indicate the ability to produce fusion proteins with saporin and cell-binding ligands for production of new reagents for further clinical and experimental use.

Original languageEnglish
Pages (from-to)6541-6548
Number of pages8
JournalJournal of Biological Chemistry
Volume268
Issue number9
Publication statusPublished - Mar 25 1993

ASJC Scopus subject areas

  • Biochemistry

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