title: Release of sphingomyelin phosphodiesterase (acid sphingomyelinase) by ammonium chloride from CL 1D mouse L-cells and human fibroblasts. Partial purification and characterization of the exported enzymes.
abstract: In cultured human fibroblasts and mouse L-cells the lysosomotropic agent, ammonium chloride, caused release of acid sphingomyelinase into the culture medium. The water-soluble enzymes were partially purified by sequential chromatography on ConA-Sepharose, octyl-Sepharose and Sepharose CL-4B. Mouse sphingomyelinase was purified up to 64-fold and human sphingomyelinase 134-fold from the culture medium. Specific activities were 925 nmol/(h X mg) and 1 434 nmol/(h X mg), respectively. The final enzyme preparations obtained were free of other lysosomal enzyme activities tested and had very similar properties: optimal activity at pH 4.8 (mouse enzyme) and pH 4.4 (human enzyme), Km values of 6.2 X 10(-5)M and 2.4 X 10(-5)M, respectively, and an apparent molecular mass of 68 kDa. In isoelectric focusing the enzymes peaked at pH 4.78 (mouse enzyme) and pH 4.75 (human enzyme).
mesh_terms: D000643:Ammonium Chloride; D000818:Animals; D055598:Chemical Phenomena; D002621:Chemistry; D002845:Chromatography / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D005347:Fibroblasts / Q000201:enzymology; D006801:Humans; D007525:Isoelectric Focusing; D007739:L Cells / Q000201:enzymology; D051379:Mice; D010727:Phosphoric Diester Hydrolases / Q000302:isolation & purification*; D013108:Sphingomyelin Phosphodiesterase / Q000302:isolation & purification*
chemical_list: D000643:Ammonium Chloride; D010727:Phosphoric Diester Hydrolases; D013108:Sphingomyelin Phosphodiesterase