Affinity Labeling of Recombinant Ricin A Chain with Procion Blue MX‐R

Recombinant ricin A chain was irreversibly modified by Procion blue MX‐R, a dichlorotriazinyl analogue of Cibacron blue F3G‐A, at pH 7.5 and 4°C in 90 h with over 95% loss of activity in an in vitro translation assay. The presence of total yeast RNA reduced the covalent attachment of Procion blue MX‐R to ricin A chain. Quantitatively modified ricin A chain contained 2 mol Procion blue MX‐R/mol 29‐kDa subunit. Tryptic digestion and resolution of the peptides by reverse‐phase high‐performance liquid chromatography yields a blue peptide corresponding to Gln5 – Arg26 of ricin A chain. Thus, a likely dye‐binding site on recombinant ricin A was identified. This region is removed from the active‐site cleft of recombinant ricin A but may be involved in its substrate binding.