Characterization of the Defective ß-Glucuronidase Activity in Canine Mucopolysaccharidosis Type VII

Abstract

Canine mucopolysaccharidosis type VII results from deficient activity of lysosomal ß-glucuronidase. Residual enzymatic activity (0.2-1.7% of normal) was detected in tissue homogenates from affected dogs. In contrast, serum and urine from affected animals had up to 15% residual activity. To further characterize the nature of the defective enzyme, hepatic ß-glucuronidase was partially purified from normal and MPS VII dogs for determination of their physical and kinetic properties. About 65% of the total ß-glucuronidase in normal canine liver required detergent for solubilization (i.e., membrane-associated), whereas only 22% of the residual activity in canine MPS VII liver was membrane-associated. Compared to the normal hepatic enzyme, the K(m) towards 4-methylumbelliferyl-ß-glucuronide was markedly increased in MPS VII dogs (i.e., 0.48 versus > 2.5 mmol/l). In contrast, the thermo-, cryo-, and pH stability properties, as well as the pH optimum (≈ 4.6), were essentially unaffected. In addition, the canine MPS VII hepatic residual activity was unresponsive to sulfhydryl reducing reagents and divalent cations, despite the fact that incubation of normal canine ß-glucuronidase with dithiothreitol and magnesium and/or calcium enhanced the activity more than 15-fold.

Effects of bone marrow transplantation on the cardiovascular abnormalities in canine mucopolysaccharidosis VII

“…4 MPS type VII (Sly syndrome, ␤-glucuronidase deficiency) was reported in mixed-breed dogs [5][6][7][8] and a colony was established at the University of Pennsylvania, School of Veterinary Medicine. All of the affected dogs are descendants of a single carrier female and were therefore homozygous for a single mutant allele, an arginine to histidine amino acid substitution at position 166.…”

Restoration of normal lysosomal function in mucopolysaccharidosis type VII cells by retroviral vector-mediated gene transfer.

“…In cultures that expressed normal amounts of GUS activity (Table 3), only -10% of the cells stained positively (Table 3), and individual cells stained more intensely than normal bone marrow cells (Fig. 2, G (42).…”

Corrective GUSB transfer to the canine mucopolysaccharidosis VII cornea using a helper-dependent canine adenovirus vector

“…Explants were from deceased donors and not usable for grafts due to endothelial density. Cornea explants were also from healthy and MPS VII dogs [12]. In vivo injections were in ~2-month old C57BL/6 mice (Charles River Laboratories), and ~6-month old gray mouse lemurs (GML, or Microcebus murinus ), a nonhuman primate that can be readily bred in captivity.…”