Fluorometric assay of neuraminidase with a sodium (4-methylumbelliferyl-alpha-D-N-acetylneuraminate) substrate. |
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| Authors: | M Potier L Mameli M Bélisle L Dallaire SB Melançon |
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| Institution: | 1. Section de Génétique Médicale, Centre de Recherche Pédiatrique, Hôpital Sainte-Justine, Montréal H3T 1CS, Québec, Canada;2. Département de Pédiatrie, Faculté de Médecine, Université de Montréal, Montréal H3T 1CS, Québec, Canada;3. Département de Biochimie, Faculté de Médecine, Université de Montréal, Montréal H3T 1CS, Québec, Canada |
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| Abstract: | This report describes the preparation of a sodium (4-methylumbelliferyl-α-d-N-acetylneuraminate) substrate and its use in a sensitive fluorometric assay of neuraminidase (EC 3.2.1.18) from Vibrio cholerae, cultured fibroblasts, and human leucocytes. V. cholerae neuraminidase showed maximum activity at pH 4.6 and an apparent Km of 1.5 mm and was activated by CaCl2 and inhibited by ethylenediaminetetraacetate, NaCl, and N-acetylneuraminic acid. The inhibition by N-acetylneuraminic acid was competitive (Ki = 6.1 mm). Cultured fibroblast and leucocyte neuraminidases showed maximum activity between pH 4.2 and 4.4 and apparent Km values of 0.13 and 0.22 mm, respectively. Neuraminidase activity was considerably reduced in cultured fibroblasts of patients with mucolipidosis types I, II, and III. |
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| Keywords: | To whom reprint requests should be sent |
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