Only sphingolipid activator protein B (SAP‐B or saposin B) stimulates the degradation of globotriaosylceramide by recombinant human lysosomal α‐galactosidase in a detergent‐free liposomal system

[摘要]

The degradation of globotriaosylceramide (GbOse₃Cer) by insect-cell derived recombinant human α-galactosidase (EC 3.2.1.22) was carried out in a detergent-free liposomal system in order to mimic intralysosomal conditions. GbOse₃Cer incorporated into unilamellar liposomes was used as the substrate, and naturally occurring sphingolipid activator proteins, rather than detergents, were used to stimulate the enzyme reaction. The degradation of GbOse₃Cer was dependent on the presence of both α-galactosidase and sphingolipid activator protein B (SAP-B or saposin B). It proceeded optimally at pH 4.6, and was enhanced by increasing amounts of both α-galactosidase (0.24–24 mU/50 μl assay) and SAP-B (0–5 μg/50 μl assay). The enzyme reaction was not affected by SAP-A, SAP-C, or SAP-D. Therefore, our results indicate that only SAP-B is essential for the degradation of GbOse₃Cer by α-galactosidase.

[学科分类] 生物化学/生物物理

[关键词] Sphingolipid activator protein B;Saposin B;α-Galactosidase;Globotriaosylceramide;Cer;ceramide or N-acylsphingosine;GbOse3Cer;globotriaosylceramide;Galα1–4Galβ1–4Glcβ1-1Cer;GM1;Galβ1–3GalNAcβ1–4Gal(3-2αNeuAc)β1–4Glcβ1-1Cer;LacCer;lactosylceramide;Galβ1–4Glcβ1-1Cer;MALDI/TOF/MS;matrix-assisted laser desorption ionization time-of-flight mass spectrometry;SAP-A;-B;-C;-D;sphingolipid activator protein A;B;C;D or saposin A;B;C;D;SAPs;sphingolipid activator proteins