Effect of immobilization on stability and properties of N-acetyl-beta-D-hexosaminidase from Turbo cornutus.

A mixture of glycosidases from the liver of the gastropod Turbo cornutus was co-immobilized with bovine serum albumin and glutaraldehyde, and then cast as membranes. The properties of immobilized N-acetyl-beta-D-hexosaminidase were studied. The recovery of N-acetyl-beta-D-hexosaminidase after immobilization was unaffected by increasing the concentration of glutaraldehyde, but was decreased by increasing the bovine serum albumin concentration. The immobilized enzyme showed enhanced resistance towards proteolytic and thermal inactivation. While the pH optimum for the soluble enzyme was 4.0, a bimodal pH curve with optima at 3.4 and 5.0 was observed after insolubilization. This bimodality was abolished when the immobilized enzyme was assayed in the presence of M NaCl. The Km values, for p-nitrophenyl 2-acetamido-2-deoxy-beta-D-glucopyranoside, of the immobilized isoenzymes of N-acetyl-beta-D-hexosaminidase were larger than those of their soluble counterparts. No loss of activity could be detected in the membrane after using it for 24 consecutive assays or after storage for at least 50 days at 4 degrees.