Human kidney α- l-Iduronidase: Purification and characterization

Abstract

α- l-Iduronidase has been purified 25,000-fold from the soluble proteins of human kidney by chromatography on heparin-Sepharose, hydroxylapatite, and Bio-Gel P-100. The α- l-iduronidase activity is associated with 80% of the protein in the most purified preparation. It has a molecular weight of 60,000 ± 6500, determined by sedimentation equilibrium, and can be dissociated by reduction into subunits of molecular weight 31,000 ± 6500 determined by polyacrylamide gel electrophoresis in sodium dodecyl sulfate in the presence of dithiothreitol. It contains glucosamine and binds to concanavalin A. The pH optimum, K m and V max for two substrates, phenyl iduronide and [ 3H]anhydromannitol iduronide, were found to be 4.0, 1.05 m m, 16 μmol/mg protein/min, and 4·5, 9 m m and 270 μmol/mg protein/min, respectively. The enzyme is of the low uptake, noncorrective form with respect to fibroblasts cultured from the skin of patients with Hurler syndrome. It is inhibited by 10 6 m p-chloromercuribenzoate and 10 −3 m Cu 2+, but is not significantly affected by other divalent cations, EDTA, or sulfhydryl compounds. Antibodies to α- l-iduronidase have been raised in goats.