Mammalian Glucocerebrosidase: Implications for Gaucher’s Disease

Glew, Robert H.; Basu, Alakananda; LaMarco, Karen L.; Prence, Elizabeth M.

doi:10.1007/978-1-4612-4502-5_1

Cited by 22 publications

(45 citation statements)

References 62 publications

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“…Glucosylceramidase is tightly bound to the lysosomal membrane; its activity is markedly impaired by delipidation and purification [4]. The purified enzyme can be reactivated by addition of acidic phospholipids, the reconstitution being facilitated by the presence of Sap C [4,111. In search for specific compounds that might have a role in the association of glucosylceramidase with membranes, we have investigated whether Sap C can promote the reconstitution of fully active glucosylceramidase by PS LUV.…”

Section: U61mentioning

confidence: 99%

“…At a concentration of PS L W up to 10 lug/ml less than 10% of the activity measured in an assay medium containing either detergents (standard assay, see section 2) or PS SUV was observed (Table I). Thus we have investigated the possibility of promoting enzyme activation by PS LUV with Sap C, a well-known glucosylceramidase activator [3,4]. As a matter of fact we have found that Sap C dramatically enhances the stimulating power of LUV (Fig.…”

Section: Effect Of Sap C On Glucosylceramidase Activation By Ps Luvmentioning

confidence: 99%

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Function of saposin C in the reconstitution of glucosylceramidase by phosphatidylserine liposomes

et al. 1993

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The function of saposin C (Sap C), a glucosylceramidase activator protein, in the enzyme stimulation by phosphatidylserine (PS) liposomes has been investigated. Using gel filtration experiments evidence was obtained for Sap C binding to PS large unilamellar vesicles (LUV) but not to glucosylceramidase. PS LUV, which by themselves are unable to tightly bind and stimulate the enzyme, acquire the capacity to also bind the enzyme after interaction with Sap C, making it express its full activity. Our results indicate that the primary step in the Sap C mode of action resides in its association with PS membranes; in turn, this association promotes the interaction between the membranes and glucosylceramidase.

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Section: U61mentioning

confidence: 99%

Section: Effect Of Sap C On Glucosylceramidase Activation By Ps Luvmentioning

confidence: 99%

Function of saposin C in the reconstitution of glucosylceramidase by phosphatidylserine liposomes

et al. 1993

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“…A defective glucosylceramidase activity leads to glucosylceramide accumulation and Gaucher disease (1)(2)(3)(4)(5). Glucosylceramidase and Sap C are highly hydrophobic proteins that become water soluble after purification; immunocytochemical studies demonstrated that a large proportions of both proteins are associated with the lysosomal membrane (6,7).…”

mentioning

confidence: 99%

“…While the function of Sap A is still debated, that of Sap C as glucosylceramidase activator is well assessed (1)(2)(3)(4)(5)13). In vitro, the Sap C-induced stimulation of glucosylceramidase requires the presence of a negatively charged phospholipid such as phosphatidylserine (PS) (5,19).…”

mentioning

confidence: 99%

“…In vitro, the Sap C-induced stimulation of glucosylceramidase requires the presence of a negatively charged phospholipid such as phosphatidylserine (PS) (5,19). In earlier studies, we provided evidence that Sap C was able to favor the association of glucosylceramidase with PS membranes and that the physical characteristics of PS vesicles markedly influenced their interaction with glucosylceramidase (20,21).…”

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Effect of Saposins A and C on the Enzymatic Hydrolysis of Liposomal Glucosylceramide

Vaccaro

Tatti

Ciaffoni

et al. 1997

Journal of Biological Chemistry

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The degradation of glucosylceramide in lysosomes is accomplished by glucosylceramidase with the assistance of, at least, another protein, saposin C (Sap C), which is generated from a large precursor together with three other similar proteins, saposins A, B, and D. In the present study, we have examined the effects of saposins on the enzymatic hydrolysis of glucosylceramide inserted in large and small phospholipid liposomes. The glucosylceramide contained in large unilamellar vesicles (LUV) was degraded by glucosylceramidase at a rate 7-8-fold lower than glucosylceramide inserted in small unilamellar vesicles (SUV). The separate addition of either Sap A or Sap C to the LUV system partially stimulated the sphingolipid degradation while saposins B and D had no effect. In the presence of both Sap A and Sap C, the rate of sphingolipid degradation was higher than the sum of the rates with the two saposins individually, indicating synergism in their actions. The stimulatory effect of the two saposins depended on the incorporation of an acidic phospholipid such as phosphatidylserine (PS) into LUV.The characteristics of glucosylceramidase activation by Sap C were different from those of Sap A. Sap C increased the rate of hydrolysis of both the artificial water soluble substrate, 4-methylumbelliferyl-␤-D-glucopyranoside, and the lipid substrate, glucosylceramide, while Sap A only stimulated degradation of the sphingolipid. Also the binding properties of Saps A and C were markedly different. At acidic pH values, Sap C bound to PS-containing LUV and promoted the association of glucosylceramidase with the membrane. In contrast, Sap A had poor affinity for the membrane even in the presence of glucosylceramide; moreover, Sap A did not potentiate the capacity of Sap C to mediate glucosylceramidase binding.In conclusion, our results show that both Sap A and Sap C are required for maximal hydrolysis of glucosylceramide inserted in PS-containing LUV, that their effects are synergistic, and that their mode of action is different. Sap C is responsible for the membrane binding of glucosylceramidase, while Sap A stimulation is possibly related to its effect on the conformation of the enzyme. It can be envisaged that Sap A in conjunction with Sap C might have a physiological role in glucosylceramide degradation.

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Use of denaturing gradient gel electrophoresis to identify mutant sequences in the β-glucocerebrosidase gene

et al. 1994

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Mammalian Glucocerebrosidase: Implications for Gaucher’s Disease

Cited by 22 publications

References 62 publications

Function of saposin C in the reconstitution of glucosylceramidase by phosphatidylserine liposomes

Function of saposin C in the reconstitution of glucosylceramidase by phosphatidylserine liposomes

Effect of Saposins A and C on the Enzymatic Hydrolysis of Liposomal Glucosylceramide

Use of denaturing gradient gel electrophoresis to identify mutant sequences in the β-glucocerebrosidase gene

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