Sanfilippo disease type B: presence of material cross reacting with antibodies against alpha-N-acetylglucosaminidase.

1. alpha-N-Acetylglucosaminidase, the enzyme deficient in Sanfilippo disease type B (mucopolysaccharidosis III B) was purified from normal human urine. An antiserum was raised in rabbits against the purified enzyme. Preincubation of the antiserum with crude alpha-N-acetylglucosaminidase from normal human urine, followed by centrifugation, led to a marked reduction of the enzyme activity in the supernatant. Formation of the antibody-enzyme complex had no influence on the activity. The thermal stability of the enzyme was markedly enhanced by complex formation with the antiserum. 2. In the urine from three patients with Sanfilippo disease type B the presence of cross-reacting material could be demonstrated by incubating the antiserum with alpha-N-acetylglucosaminidase in the presence of Sanfilippo B urine or by pretreatment of the antiserum with Sanfilippo B urine. 3. Immunodiffusion and immunoelectrophoresis of crude normal or Sanfilippo B urine gave rise to up to four precipitation lines, only one of which exhibited alpha-N-acetylglucosaminidase activity in the case of normal urine. Purified alpha-N-acetylglucosaminidase yielded only a single precipitation line. After adsorption with the purified enzyme the antiserum did not cross react with any of the urinary proteins. 4. On a quantitative determination of cross-reacting material using Sepharose immobilized antibodies in the urine from two Sanfilippo B patients the amount of cross-reacting material appeared to be less than one fourth of the amount of alpha-N-acetylglucosaminidase protein in an age-matched control urine. The cross-reacting material present in the urine of Sanfilippo B patients had a significant lower binding affinity for antibodies against alpha-N-acetylglucosaminidase than preparations from normal human urine. Taking into account this lower binding affinity, it can be calculated that the amount of cross-reacting material in the urine of Salfilippo B patients exceeds that of normal controls. 5. It is concluded that Sanfilippo disease type B is due to a mutation of a structural gene coding for alpha-N-acetylglucosaminidase. The mutation affects the catalytical and immunological properties of the enzyme protein.