Biospecific Elution from Non-Specific Adsorbents (‘Affinity Elution’)

Enzymes adsorbed to ion exchangers are in many cases specifically desorbed by interaction with their ligands. This phenomenon has been variously called ‘specific elution with substrate’ or ‘affinity elution’ in the literature (von der Haar, 1974). Because the most effective ligands for elution are those containing charged groups identical with those on the ion exchanger (positive groups in the case of anion exchangers, negative groups in the case of cation exchangers), it has been assumed, that desorption results from differences between the net charge of the free enzyme and the enzyme-ligand complex. Theoretically, however, even a neutral ligand may lead to desorption of an enzyme, if the charged groups responsible for adsorption are located in the binding site for the ligand and are sterically shielded from interaction with the ion exchanger after complex formation or if a conformational change occurs that prevents charged groups from interacting with the exchanger. Besides charge differences between the enzyme and the enzyme-ligand complex, other parameters influencing the effectiveness of elution are the strength of binding of the enzyme to the ion exchanger and the affinity between enzyme and ligand. An equilibrium exists between enzyme (E) bound to the ion exchanger (IE) (eqn. 1) and the enzyme bound to the ligand (lig) in solution (eqn. 2).