Isolation of high-specific-activity subunits of cholera toxin by reversed-phase high-performance liquid chromatography

Facile, rapid procedures for the separation of active cholera toxin subunits were developed, based on high-performance liquid chromatography (HPLC) with a Nucleosil C₈ reversed-phase column. These procedures were capable of completely resolving subunits A and B as well as S-carboxymethylated or reduced α-, γ-, and β-chains. The binding of HPLC-purified B subunit to GM₁ ganglioside was essentially identical to that of cholera toxin when compared on a molar basis. The adenosine 5′-diphosphate-ribosyltransferase activity of HPLC-purified A subunit, reduced α-chain, or carboxymethyl α-chain was also determined to be reasonably high compared to that of cholera toxin or commercially prepared A subunit.