PURIFICATION OF RECOMBINANT RICIN-A CHAIN WITH IMMOBILIZED TRIAZINE DYES

Immunotoxins, such as those based on ricin A chain, must be rigorously purified before they can be administered in vivo. The work described in this paper investigates the interaction between recombinant ricin A chain and several triazine dyes and other ligands that may be of value in its purification. All ligands displayed a high affinity (dissociation constants 3-20 ELM) and are displaced from their binding sites on the protein by polynucleotides, heparin and synthetic polyphosphates, but not by mono- or dinucleotides. Affinity chromatography on the immobilised dyes, Procion Red H-3B, Procion Red HE-3B, Procion Red HE-7B and Procion Yellow HE-4R, resulted in a one-step purification of recombinant ricin A chain from an Escherichia coli fermentation extract to 94-98% purity and with a > 95% yield. These materials are far superior to purification on the conventional dye, Cibacron Blue F-3GA, and show promise for the isolation of immunotoxins from immunoconjugation mixtures.