Human mucopolysaccharidosis VII (MPS VII, Sly syndrome) results from a deficiency of -glucuronidase (GUS) and has been associated with a wide range in severity of clinical manifestations. To study missense mutant models of murine MPS VII with phenotypes of varying severity, we used targeted mutagenesis to produce E536A and E536Q, corresponding to active-site nucleophile replacements E540A and E540Q in human GUS, and L175F, corresponding to the most common human mutation, L176F. The E536A mouse had no GUS activity in any tissue and displayed a severe phenotype like that of the originally described MPS VII mice carrying a deletion mutation (gus mps/mps ). E536Q and L175F mice had low levels of residual activity and milder phenotypes. All three mutant MPS models showed progressive lysosomal storage in many tissues but had different rates of accumulation. The amount of urinary glycosaminoglycan excretion paralleled the clinical severity, with urinary glycosaminoglycans remarkably higher in E536A mice than in E536Q or L175F mice. Molecular analysis showed that the Gus mRNA levels were quantitatively similar in the three mutant mouse strains and normal mice. These mouse models, which mimic different clinical phenotypes of human MPS VII, should be useful in studying pathogenesis and also provide useful models for studying enzyme replacement therapy and targeted correction of missense mutations.knock-in mice ͉ point mutation ͉ Cre-loxP M ucopolysaccharidosis type VII (MPS VII or Sly syndrome) is a mucopolysaccharide storage disease resulting from a deficiency of -glucuronidase (GUS, EC 3.2.1.31) (1). In MPS VII, chondroitin sulfate, dermatan sulfate, and heparan sulfate are only partially degraded and accumulate in the lysosomes of many tissues, leading to cellular and organ dysfunction. MPS VII has also been reported in canine, murine, and feline species. Human patients with MPS VII display a wide range of clinical severity, and Ͼ45 different mutations have been found in the GUS gene (2-13). Around 90% of these are point mutations. L176F accounts for Ϸ20% of mutant alleles. This mutation was first identified in a Mennonite family (7), and then observed in other populations. Most patients homozygous for L176F have a mild phenotype. This mutation is interesting because cells from L176F patients have Ͻ1% of normal GUS activity, but expression of the L176F cDNA in COS cells produced nearly as much enzyme activity as the WT control cDNA (7).Characterization of human GUS protein by x-ray crystallography and homology comparisons among several species suggested R382, E451, and E540 as active-site residues (14). E540 was identified as the active-site nucleophile of the human enzyme (15, 16). Recombinant E540A human GUS had no catalytic activity, but the E540Q GUS showed 0.3% of WT activity (16). One severely affected MPS VII patient with a null mutation at this residue, E540K, has been identified (S.T. and W.S.S., unpublished observation).The original MPS VII (gus mps/mps ) mice with a 1-bp deletion in exon 10 have morphologi...