Radioassay method of neuraminidase towards N-acetylneuraminosyl hexasaccharides.
We devised a sensitive method to assay for neuraminidase activities towards alpha-(2 leads to 3)-N-acetylneuraminosyl hexasaccharide and alpha-(2 leads to 6)-N-acetylneuraminosyl hexasaccharide, which were isolated from the urine of a patient with adult sialidosis with partial deficiency of beta-galactosidase. Standard assay conditions for the determination of these neuraminidase activities were established and the radiolabeled reduced derivatives of these substrates were used. The fibroblast neuraminidase had its maximum activity at pH 4.0-4.2, with Km values of 2.22 X 10(-3) and 4.17 X 10(-3) mol/l and Vmax values of 76.9 and 28.6 nmol . mg-1 protein . h-1 towards the 2 leads to 3 isomer and the 2 leads to 6 isomer, respectively. Neuraminidase deficiencies were found in the fibroblasts of adult sialidosis, mucolipidosis II and III. These studies were compared with the neuraminidase activity towards alpha-(2 leads to 3)-N-acetylneuraminosyl lactose.
D002239:Carbohydrate Metabolism, Inborn Errors / Q000201:enzymology; D005347:Fibroblasts / Q000201:enzymology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007785:Lactose / Q000031:analogs & derivatives* / Q000378:metabolism; D007787:Lactose Intolerance / Q000201:enzymology; D009439:Neuraminidase / Q000378:metabolism*; D009844:Oligosaccharides / Q000378:metabolism*; D012794:Sialic Acids / Q000378:metabolism*
D009844:Oligosaccharides; D012794:Sialic Acids; C000020:N-acetylneuraminoyllactose; D009439:Neuraminidase; D007785:Lactose