Sphingolipid Activator Protein D (sap-D) Stimulates the Lysosomal Degradation of Ceramide in Vivo

Klein, Andreas; Henseler, Margarete; Klein, Christian; Suzuki, Kazuyuki; Harzer, K.; Sandhoff, Konrad

doi:10.1006/bbrc.1994.1612

Cited by 153 publications

(97 citation statements)

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“…Previous data obtained by computer analysis and comparison of the secondary structures of sphingolipid activator proteins and ASM suggest a high similarity between the ASM domain N-terminal to the hinge region and the activator proteins (Ponting, 1994). Since ASM, in contrast to most other lysosomal hydrolases degrading sphingolipids with short hydrophilic groups, does not need stimulation by known sphingolipid activator proteins in vivo (Klein et al, 1994), the mature enzyme of 70 kDa may already contain an intrinsic sphingomyelin activating domain loosely attached to the enzymatic domain (57 kDa) by a proline spacer.…”

Section: Phosphorylation Of the Asm Oligosaccharidesmentioning

confidence: 90%

Functional Characterization of the N‐glycosylation Sites of Human Acid Sphingomyelinase by Site‐Directed Mutagenesis

Ferlinz¹,

Hurwitz²,

Moczall³

et al. 1997

European Journal of Biochemistry

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Most soluble lysosomal enzymes require a mannose-6-phosphate recognition marker present on asparagine-linked oligosaccharides for proper targeting to lysosomes. We have determined the influence of the six potential N-linked oligosaccharide chains of human acid sphingomyelinase (ASM) on catalytic activity, targeting, and processing of the enzyme. Each N-glycosylation site was modified by site-directed mutagenesis and subsequently expressed in COS-1 cells. Evidence is presented that five of these sites are used. Elimination of the four N-terminal glycosylation sites does not disturb lysosomal targeting, processing, or enzymatic activity. However, removal of the two C-terminal N-glycosylation sites inhibits the formation of mature enzyme. Absence of glycosylation site five resulted in rapid cleavage of the primary translation product to an enzymatically inactive protein which accumulated inside the endoplasmic reticuIudGolgi, whereas deletion of glycosylation site six led to the formation of an inactive ASM precursor, also retained inside the endoplasmic reticuludGolgi. Our results also provide evidence that the site of early proteolytic cleavage of newly synthesized ASM must be located between the second and third glycosylation sites.

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Section: Phosphorylation Of the Asm Oligosaccharidesmentioning

confidence: 90%

Functional Characterization of the N‐glycosylation Sites of Human Acid Sphingomyelinase by Site‐Directed Mutagenesis

Ferlinz¹,

Hurwitz²,

Moczall³

et al. 1997

European Journal of Biochemistry

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“…Sap D has been proposed to have a role in ceramide degradation, since its addition to the culture medium of fibroblasts from a patient suffering from prosaposin deficiency resulted in the reduction of ceramide storage (22). Recently it has been .…”

Section: Discussionmentioning

confidence: 99%

Saposin D Solubilizes Anionic Phospholipid-containing Membranes

Ciaffoni

Salvioli

Tatti

et al. 2001

Journal of Biological Chemistry

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“…Control radioactivity was measured by adding SAPs and enzyme only after termination of the incubation. SAPs, SAP-A to D, were purified from human tissues [7]. ct-Galactosidase activity was measured with 4-methylumbeUiferyl-eto-galactopyranoside as the artificial substrate [10].…”

Section: Enzyme Assaysmentioning

confidence: 99%

“…Four of the five known SAPs, SAP-A to D, also called saposin A-D, are derived from a common precursor [4][5][6]. Despite their homology, like the identical locations of cysteine residues and glycosylation sites, SAP-B, C and D have different functions, which comprise lipid binding and enzyme stimulating specificities [3,7]. On the other hand, the physiological function of SAP-A remains unclear because no SAP-A deficiency has been characterized so far [3].…”

Section: Introductionmentioning

confidence: 99%

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

et al. 1996

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The degradation of globotriaosylceramide (GbOse3Cer) by insect-cell derived recombinant human ct-galactosidase (EC 3.2.1.22) was carried out in a detergent-free iiposomal 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 ct-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 ctgalactosidase (0.24-24 mU/50 lal assay) and SAP-B (0-5 pg/50 p] 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 ctgalactosidase.

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Sphingolipid Activator Protein D (sap-D) Stimulates the Lysosomal Degradation of Ceramide in Vivo

Cited by 153 publications

References 0 publications

Functional Characterization of the N‐glycosylation Sites of Human Acid Sphingomyelinase by Site‐Directed Mutagenesis

Functional Characterization of the N‐glycosylation Sites of Human Acid Sphingomyelinase by Site‐Directed Mutagenesis

Saposin D Solubilizes Anionic Phospholipid-containing Membranes

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

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