Cation-exchange chromatography of Clostridium botulinum type a toxin on amberlite IRC-50 resin at pH 5.55

Abstract

The hemagglutinin-free toxin (the α fraction) isolated from crystalline toxin of Clostridium botulinum type A, homogeneous by several criteria, was examined on carboxymethyl-Sephadex, sulfoethyl-Sephadex and Amberlite IRC-50 (XE-64) columns to attempt further resolution and to provide additional tests of its homogeneity. Satisfactory conditions for chromatography were obtained only with Amberlite IRC-50 (XE-64) resin. As an analytical tool this resin proved more sensitive than a DEAE-cellulose column since it was able to separate the hemagglutinin when present as trace contaminant in the α fraction, which could not be achieved by anion-exchange chromatography. Although the IRC-50 column separated the 150 000 mol. wt. specie of the α fraction from its aggregated forms, the toxin could not be fractionated into subunits. These observations were consistent with the result of gel filtration of the α fraction and its aggregates on Sephadex columns. The present report also demonstrates for the first time that a protein as large as 150 000 mol. wt. can be successfully chromatographed on IRC-50 (XE-64) resin at pH 5.55.