1. 1. A crude preparation of membrane-bound neuraminidase (mucopolysaccharide N-acetylneuraminylhydrolase, EC 188.8.131.52), obtained from calf brain, was used. This preparation was free from the soluble and lysosomal neuraminidases and depleted of endogenous substrates. 2. 2. The enzyme showed: (a) optimum pH 4.0 for gangliosides GD1a, GD1b, GT1b and GQ1; 3.8 for ovine submaxillary mucin, ovine submaxillary mucin-sialoglycopeptides and brain sialoglycopeptides; 3.1 for sialyllactose (C-3 isomer); (b) apparent higher affinity for gangliosides (Km of the order 10-5 M) than for sialyllactose (Km 0.68·10-3 M), ovine submaxillary mucin and sialoglycopeptides; (c) higher V for gangliosides (1.26-2.12 units/mg protein), lower for sialyllactose (1.18 units/mg protein) and sialoglycoprotein substrates (0.27-0.41 units/mg protein); (d) inhibition by excess substrate (over 0.15-0.2 mM) only in the case of gangliosides; (e) maximum rate of hydrolysis of gangliosides at 70 °C (24 units/mg protein in the case of ganglioside GD1a); (f) considerable stability. 3. 3. Na+ and Li+ did not influence the enzyme activity; K+ activated below 0.1 M; NH4 + started inhibiting at 0.01 M. All bivalent cations tested inhibited the enzyme: Hg2+ from 10-6 M, Cu2+ from 10-5 M, Ca2+ from 10-3 M. Anions had no appreciable influence on the enzyme activity, at concentrations up to 5·10-2 M.
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