FELINE MUCOPOLYSACCHARIDOSIS TYPE-VI - CHARACTERIZATION OF RECOMBINANT N-ACETYLGALACTOSAMINE 4-SULFATASE AND IDENTIFICATION OF A MUTATION CAUSING THE DISEASE

Mucopolysaccharidosis type VI (RIPS VI) is an autosomal recessive dise ase caused by a deficiency of N-acetylgalactosamine 4-sulfatase (4S) l eading to the lysosomal accumulation and urinary excretion of dermatan sulfate, RIPS VI has also been described in the Siamese cat, As an in itial step toward enzyme replacement therapy with recombinant feline 4 S (rf4S) in RIPS VI cats, the feline 4S cDNA was isolated and expresse d in CHO-KI cells and rf4S was immunopurified from the culture medium. SDS-polyacrylamide gel electrophoresis analysis showed that the precu rsor form of immunopurified rf4S was a 66-kDa polypeptide that underwe nt maturation to a 43-44-kDa polypeptide. Endocytosis of rf4S by cultu red feline RIPS VI myoblasts was predominantly mediated by a mannose g -phosphate receptor and resulted in the correction of dermatan sulfate storage, The mutation causing feline MPS VI was identified as a base substitution at codon 476, altering a leucine codon to a proline (L476 P), The L476P allele displayed no detectable 45 activity when expresse d in CHO-KI. cells and was observed only as a ''precursor'' polypeptid e that was not secreted into the medium, Identification of the mutatio n has allowed the development of a rapid PCR-based screening method to genotype individuals within the cat colony.