Human alpha-L-iduronidase. II. Comparative biochemical and immunologic properties of the purified low and high uptake forms.
The physicokinetic and immunologic properties of the purified low and high uptake forms of the human lysosomal hydrolase, alpha-L-iduronidase, have been determined and compared. The apparent Km and Vmax values for the low and high uptake forms were similar toward two artificial substrates, 4-methylumbelliferyl-alpha-L-iduronide (0.07 and 0.06 mmol/l; 16.15 and 14.85 mumol/min/mg, respectively), and phenyl-alpha-L-iduronide (1.42 and 1.66 mmol/l; 0.83 and 1.05 mumol/min/mg, respectively), and one natural substrate, anhydro-[3H]-mannitol-iduronide (0.86 and 1.04 mmol/l; 2.50 and 2.79 mumol/min/mg, respectively). The pH optima for both purified forms also were similar for each of the three substrates ( approximately 3.50, approximately 3.50, and approximately 4.50, respectively). Heparin markedly inhibited the 4-methylumbelliferyl-alpha-L-iduronide activities of both the low and high uptake forms, while dermatan sulfate and heparan sulfate were more inhibitory toward the low uptake activity. EDTA was a potent inhibitor of both enzyme forms; the divalent cations, Mg2+ and Ca2+, could recover up to 30% of the enzymatic activities after EDTA treatment. p-Chloromercuribenzoate and maleate also were inhibitory, whereas dithiothreitol and 2-mercaptoethanol were stimulatory. Both enzyme forms had similar thermostabilities ; the half-lives at 45, 52, and 60 degrees C were about 38, 24 and 12 min, respectively. The low and high uptake forms were immunologically cross-reactive as demonstrated by Ouchterlony double immunodiffusion and immunotitration studies using anti-human low uptake antibodies.
D004705:Endocytosis; D004789:Enzyme Activation; D006025:Glycosaminoglycans / Q000494:pharmacology; D006026:Glycoside Hydrolases / Q000378:metabolism*; D006358:Hot Temperature; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007068:Iduronidase / Q000037:antagonists & inhibitors / Q000276:immunology / Q000378:metabolism*; D007700:Kinetics; D008168:Lung / Q000201:enzymology
D006025:Glycosaminoglycans; D006026:Glycoside Hydrolases; D007068:Iduronidase