ABSTRACT
Two types of ribosome-inactivating proteins have been purified from plants: type I, which is made of a single polypeptide chain, and type II, which consists of two nonhomologous subunits. This chapter describes some of the properties of these proteins, named Luffa ribosome-inactivating proteins, and summarizes the cloning and expression of a related protein. A variety of single-chain ribosome-inactivating proteins have been isolated from plants and chemically linked to antibodies to form highly toxic, specific immunotoxins. Amino acid sequences were determined in a Beckman automated peptide sequencer. The fusion protein with ß-galactosidase did not show significant inhibition of protein synthesis in a cell-free system. Plaques showing a positive signal were then re-purified by plating at a lower density until all plaques in the dish were positive for the expression of fusion protein. In our cloning procedure, four rounds of plaque purification were necessary to reach that stage.
