A fluorometric assay using 4-methylumbelliferyl alpha-L-iduronide for the estimation of alpha-L-iduronidase activity and the detection of Hurler and Scheie syndromes.
Incubation of 4-methylumbelliferyl alpha-L-iduronide with whole cell homogenates prepared from cultured skin fibroblasts and amniotic cells, and peripheral blood leukocytes gave 4-methylumbelliferone which was easily measured fluorometrically. This reaction, presumably due to the action of alpha-L-iduronidase, has a maximum hydrolytic activity at pH 3.25. The apparent KM value of alpha-L-iduronidase in leukocyte whole cell homogenates for this substrate was 179 mumol/l compared to 353, 41 and 166 mumol/l for other alpha-L-iduronidase substrates phenyl alpha-L-iduronide, iduronosyl anhydrol[1-3H]mannitol 6-sulfate and iduronosyl anhydro[1-3H]mannitol respectively; the corresponding Vmax values were 617, 394, 158 and 10 pmol/min/mg protein respectively. Incubation of the 4-methylumbelliferyl alpha-L-iduronide with whole cell homogenates prepared from cultured skin fibroblasts and leukocytes from a Hurler patient gave 4-methylumbelliferone at a rate more than 20 times less than found for control normal preparations. 4-Methylumbelliferyl alpha-L-iduronide is a sensitive, convenient and superior substrate to phenyl alpha-L-iduronide for the assay of alpha-L-iduronidase activity, but is not a suitable replacement for the radiolabelled substrate iduronosyl anhydro[1-3H]mannitol 6-sulfate.
D000653:Amniotic Fluid / Q000166:cytology / Q000201:enzymology; D005470:Fluorometry / Q000379:methods*; D006026:Glycoside Hydrolases / Q000032:analysis*; D006801:Humans; D007068:Iduronidase / Q000032:analysis*; D007962:Leukocytes / Q000201:enzymology; D009083:Mucopolysaccharidoses / Q000175:diagnosis*; D008059:Mucopolysaccharidosis I / Q000175:diagnosis* / Q000201:enzymology; D012867:Skin / Q000166:cytology / Q000201:enzymology; D014468:Umbelliferones
D014468:Umbelliferones; D006026:Glycoside Hydrolases; D007068:Iduronidase