[55] Protein activator (coglucosidase) for the hydrolysis of β-glucosides

Radin, Norman S.; Berent, Susan L.

doi:10.1016/0076-6879(82)83057-7

Cited by 16 publications

(6 citation statements)

References 11 publications

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“…Isolation of human cohydrolase. The previously described method was used [4,5] but the elution conditions had to be modified for the hydroxyapatite column (see Results). One control spleen was removed surgically from a patient with pancreatic carcinoma, another from a patient with idiopathic thrombocytopenic purpura.…”

Section: Methodsmentioning

confidence: 99%

“…with a stock preparation of glucosidase, using methylumbelliferyl glucoside as substrate [4,5]. One activity unit corresponds to a 50% stimulation.…”

Section: Cohydrolase Assay the Activator Was Measuredmentioning

confidence: 99%

“…The preparation also stimulated the hydrolysis of the naturally occurring fl-glucoside, glucosyl ceramide [3]. We have obtained a highly purified material having similar properties from normal bovine spleen [4,5]. Later work, with this and a preparation from human spleen, showed that it can also stimulate the sphingolipid hydrolases, cerebroside fl-galactosidase and sphingomyelinase [6,7].…”

Section: Introductionmentioning

confidence: 96%

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The cohydrolases in human spleen that stimulate glucosyl ceramide β-glucosidase

Berent

Radin

1983

Biochimica et Biophysica Acta (BBA) - Protein Structure and Mol

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A family of beta-glucosidase-stimulating proteins (called cohydrolase SPH-I here) was isolated from bovine, Gaucher human and control human spleens. All preparations exhibited a similar pattern of four major electrophoretic bands in polyacrylamide when stained with the cationic dye, Stains-All. The bovine bands migrated more rapidly, while the two types of human cohydrolase migrated very similarly. The two human preparations differed in several respects: the concentration was much higher in Gaucher spleen; the Gaucher factors eluted a little earlier from gel permeation and decyl agarose columns; much more of the cohydrolase was bound by a concanavalin A column; the control bands stained less intensely in gels than the Gaucher bands. Antibodies raised in rabbits to bovine cohydrolase reacted with all three preparations. All four bands from Gaucher cohydrolase showed similar ability to stimulate glucosidase and to bind the antibodies. It is evident that the cohydrolases from control and Gaucher spleens are similar in many respects, yet differ in some secondary fashion, possibly in carbohydrate content. It is suggested that Gaucher cohydrolase is formed from normal cohydrolase by the nonenzymatic action of cellular glucose over a period of many years, due to slowed catabolism of the cofactor.

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Section: Methodsmentioning

confidence: 99%

“…with a stock preparation of glucosidase, using methylumbelliferyl glucoside as substrate [4,5]. One activity unit corresponds to a 50% stimulation.…”

Section: Cohydrolase Assay the Activator Was Measuredmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 96%

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The cohydrolases in human spleen that stimulate glucosyl ceramide β-glucosidase

Berent

Radin

1983

Biochimica et Biophysica Acta (BBA) - Protein Structure and Mol

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“…SAP-2 Assay-Samples were assayed for stimulatory activity toward a partially purified preparation of glucosidase by incubation with methylumbelliferyl glucoside in acetate buffer, pH 4.5, with Triton X-100 as previously described (12).…”

Section: Methodsmentioning

confidence: 99%

The carbohydrate moiety of the activator protein for glucosylceramide β-glucosidase

Sano¹,

Radin²

1988

Biochemical and Biophysical Research Communications

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SAP-2 is a family of heat-stable, acidic glycoproteins which stimulate enzymatic hydrolysis of glucosylceramide. We studied the carbohydrate moieties of a ConA-binding form of SAP-2. The protein contained glucosamine, galactose, mannose, and fucose; galactosamine and sialic acid were not detectable. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and silver staining showed three bands of 6.5, 8.5, and 10 kDa. After deglycosylation with peptide N-glycosidase, SAP-2 eluted more slowly from the C4 column and showed a single band of 4 kDa. From carbohydrate analysis it was evident that deglycosylation had removed more than 90% of the sugars. These data indicate that SAP-2 possesses N-linked complex or hybrid type oligosaccharide chains. The specific activity of the deglycosylated protein in the glucosidase stimulation assay was unaffected.

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“…First, the strategy for treating Gaucher disease, substrate reduction through the targeting of glucosylceramide synthesis, was grounded in a long-standing effort to better understand glycosphingolipid metabolism. This work included the elucidation of techniques for the isolation of cerebrosides (19), the characterization of distinct pathways for glucosylceramide and galactosylceramide synthesis and degradation (15), and the discovery of cofactors that regulated these pathways (18). A detailed understanding of the intended target for inhibition of glycosphingolipid synthesis was invaluable in the identification of potential lead compounds and the determination of whether such compounds had both activity and specificity.…”

Section: Lessons Learnedmentioning

confidence: 99%