ImmobilizedClostridium perfringensNeuraminidase. Substrate Cleavage and Enzyme Release During Incubation

Abstract

Pure C. perfringens neuraminidase [EC 3.2.1.18] was immobilized on Sepharose 4B, azido-Sepharose 4B and controlled pore glass (CPG)-glycophase using different coupling procedures. The immobilized enzyme showed increased stability under various conditions relative to the soluble enzyme. The low release of active enzyme from the supports under incubation conditions was quantitated using a highly sensitive radioactive assay. The activity of the immobilized enzyme was dependent on the nature of the support and the substrate. Activity decreased with increasing substrate MW, but the enzyme showed improved cleavage with GD1a micelles and human erythrocytes, substrates having ordered surface properties. Uses of immobilized neuraminidase in biochemistry and cell biology are considered and evaluated relative to the measured release of enzyme from the supports reported and to the molecular size and organization of possible substrates.