title: Human eosinophil arylsulfatase B. Structure and activity of the purified tetrameric lysosomal hydrolase.
abstract: Arylsulfatase B from human eosinophils was purified free of contaminating proteins by gel filtration and sequential affinity chromatography on Affi-Gel Blue and zinc chelate Sepharose. 50 micrograms of the purified enzyme presented as a single stained band on alkaline disc gel electrophoresis. In both goats and rabbits, the purified enzyme elicited monospecific antisera that yielded single precipitation arcs on Ouchterlony analysis with a human eosinophil extract and the purified enzyme; the immunoprecipitation lines fused in a pattern of identity, providing immunochemical evidence for the homogeneity of the purified enzyme. On sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis, a dominant lower molecular weight protein and three other bands with molecular weights approximately two, three, and four times that of the major protein band were resolved. The prominence of the less rapidly migrating protein bands increased relative to the major band if the enzyme was maintained under acidic conditions or was reacted with the cross-linking agent dimethyl suberimidate under alkaline conditions before SDS-polyacrylamide gel electrophoresis, supporting the conclusion that the enzyme consists of four subunits. Two stained bands were present on acid disc gel electrophoresis; they were composed of oligomeric forms of enzyme on analysis by SDS-polyacrylamide gel electrophoresis in a second dimension. A minimum molecular weight of 70,190 was determined from amino acid composition analysis for the tetrameric form of the enzyme. The specific functional activity of the purified arylsulfatase B was concentration and time dependent, compatible with its association or dissociation into subunit forms with differing specific activities. Factors that govern subunit interactions of arylsulfatase B, including local enzyme concentration and pH, provide mechanisms for regulating the enzymatic activity of this lysosomal hydrolase.
mesh_terms: D002803:Chondro-4-Sulfatase / Q000097:blood* / Q000302:isolation & purification; D004804:Eosinophils / Q000201:enzymology*; D006801:Humans; D008247:Lysosomes / Q000201:enzymology; D046911:Macromolecular Substances; D011485:Protein Binding; D011487:Protein Conformation; D013429:Sulfatases / Q000097:blood*
chemical_list: D046911:Macromolecular Substances; D013429:Sulfatases; D002803:Chondro-4-Sulfatase