An endogenous activator protein in human placenta for enzymatic degradation of glucosylceramide.
An endogenous, heat-stable and pronase-sensitive activator for enzymatic hydrolysis of glucosylceramide was detected in the crude lysosome-mitochondria fraction of human placenta. Its properties differ distinctly in several important respects from those of the previously described glucosylceramidase activator. The activator reported here had no effect on crude glucosylceramidase with either glucosylceramide or 4-methylumbelliferyl-beta-D-glucopyranoside as the substrate in the presence of either sodium taurocholate or phosphatidylserine. On the contrary, glucosylceramide hydrolysis by the enzyme partially purified through Octyl-Sepharose 4B chromatography was stimulated by this activator 6-9-fold in the presence of either sodium taurocholate or phosphatidylserine. The Km for glucosylceramide in the presence of the activator was 1/3 of that without the activator. In the crude enzyme fraction, the activator was present in a 16-fold excess over the minimum amount necessary for full activation of the enzyme. Hydrolysis of the fluorogenic substrate by the post-Octyl-Sepharose enzyme, however, was not stimulated by the activator. Similarly, hydrolysis of galactosylceramide by galactosylceramidase obtained from the same Octyl-Sepharose chromatography was not stimulated. Our observations are consistent with the idea that glucosylceramidase is saturated by, or perhaps tightly associated with, this activator in the placenta and that they are dissociated by the Octyl-Sepharose chromatography. In fact, the properties of the combined post-Octyl-Sepharose enzyme and activator closely mimic those of the crude enzyme without added activator.
D002847:Chromatography, Agarose; D004789:Enzyme Activation; D005260:Female; D005959:Glucosidases / Q000378:metabolism*; D005962:Glucosylceramidase / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D008247:Lysosomes / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism; D010920:Placenta / Q000201:enzymology / Q000378:metabolism*; D011257:Pregnancy Proteins / Q000302:isolation & purification* / Q000502:physiology; D012685:Sepharose / Q000031:analogs & derivatives
D011257:Pregnancy Proteins; C027176:octyl-sepharose CL-4B; D012685:Sepharose; D005959:Glucosidases; D005962:Glucosylceramidase