Cloning and expression of the gene involved in Sanfilippo B syndrome (mucopolysaccharidosis III B)

Sanfilippo B syndrome is caused by a deficiency of alpha-N-acetylglucosaminidase, a lysosomal enzyme involved in the degradation ofheparan sulphate. Accumulation of the substrate in lysosomes results indegeneration of the central nervous system with progressive dementia oftencombined with hyperactivity and aggressive behaviour. In order to clone thedeficient gene, we purified the enzyme from human placenta and obtainedamino acid sequence information. Alignment of one of the CNBr generatedinternal peptides to sequence from the database revealed the chromosomallocation of the gene in the 5' upstream flanking region of the gene for17-beta-hydroxysteroid-dehydrogenase at 17q21.1. The available DNA sequencewas used to clone the cDNA coding for alpha-N- acetylglucosaminidase andanalyse its gene structure. The gene is fully contained in the 5' upstreamflanking region of the gene for 17-beta- hydroxysteroid-dehydrogenase andinterrupted by five introns. The cDNA clone has a length of 2575 bp andencodes a protein of 743 amino acids. Chinese hamster ovary cellstransfected with the cDNA construct show alpha-N-acetylglucosaminidaseactivity about 17-fold over background. This will allow correction studieswith NAG deficient Sanfilippo B cell lines and facilitate the developmentof enzyme replacement therapy for these patients.