The chromatography of enzymes on group-specific affinity adsorbents, such as nucleotides, has been used to optimize the design of biospecific adsorbents (1). Despite the widespread application of these columns to the purification of many nucleotide-dependent enzymes, one drawback is apparent. The operational capacities of these columns are frequently low; 0.1 to 0.5% of the total ligand is common. It has been suggested that this phenomenon relates to the association constant (2). Some other small ligands show increased operational capacities when compared with nucleotides, for example, immobilized Cibacron Blue (3). Cibacron Blue-Sepharose in our hands is more specific for NAD+-dependent enzymes than is suggested by the literature. On the other hand we havefound that another triazine dye, Procion Red HE3B, also shows high operational capacities and appears to be specific for NADP+-dependent dehydrogenases.
KeywordsAffinity Chromatography Association Constant Operational Capacity Dependent Enzyme Small Ligand
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