BIOLOGICALLY-ACTIVE RECOMBINANT RICIN-B CHAIN PRODUCED IN ESCHERICHIA-COLI

A recombinant B subunit (rRTB) of ricin, a plant toxin, was obtained. The heterologous protein synthesized in Escherichia coli formed inclus ion bodies. Denaturation and renaturation of the protein in the presen ce of glutathione and lactose yielded a soluble, biologically active a nd stable product. The rRTB had no toxic effect on HUT102 cells (ID50 > 10(-6) M). Antigenic properties and the galactose-binding activity w ere the same for both natural and recombinant RTB; rRTB, however, prov ed to be more sensitive to trypsin. A chimeric toxin (rRTB-RTA) was re constructed using rRTB and the natural ricin A subunit. The cytotoxic activity of the rRTB-RTA heterodimer toward HUT102 cells was one-tenth that of the natural RTB-RTA heterodimer and ricin. These results sugg est that carbohydrate chains bound to the RTB molecules are required t o maintain the protein in a stable form and to prevent proteolysis of the protein in intracellular vesicles.