Human Placental Sialidase: Further Purification and Characterization

Hiraiwa, Masao; Nishizawa, Masatoyo; Uda, Yutaka; Nakajima, Takashi; Miyatake, Tadashi

doi:10.1093/oxfordjournals.jbchem.a122245

Cited by 40 publications

(27 citation statements)

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“…Few reports on human lysosomal neuraminidase have been published, because the enzyme has low stability during purification and requires PPCA for the expression of its activity (Verheijen et al 1982(Verheijen et al , 1985(Verheijen et al , 1987Hiraiwa et al 1988;van der Horst et al 1989). Recent cDNA cloning for human lysosomal neuraminidase has revealed that it encodes a 45-kDa protein with three potential N-linked glycosylation sites (Bonten et al 1996;Pshezhetsky et al 1997;Milner et al 1997).…”

Section: Discussionmentioning

confidence: 99%

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Molecular and structural studies of Japanese patients with sialidosis type 1

et al. 2000

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To gain insight into the pathogenesis of sialidosis type 1, we performed molecular investigations of two unrelated Japanese patients. Both of them are compound heterozygotes for base substitutions of 649 G-to-A and 727 G-to-A, which result in amino acid alterations V217M and G243R, respectively. Using homology modeling, the structure of human lysosomal neuraminidase was constructed and the structural changes caused by these missense mutations were deduced. The predicted change due to V217M was smaller than that caused by G243R, the latter resulting in a drastic, widespread alteration. The overexpressed gene products containing these mutations had the same molecular weight as that of the wild type, although the amounts of the products were moderately decreased. A biochemical study demonstrated that the expressed neuraminidase containing a V217M mutation was partly transported to lysosomes and showed residual enzyme activity, although a G243R mutant was retained in the endoplasmic reticulum/Golgi area and had completely lost the enzyme activity. Considering the data, we surmise that the V217M substitution may be closely associated with the phenotype of sialidosis type 1 with a late onset and moderate clinical course.

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Section: Discussionmentioning

confidence: 99%

“…1. 18) is an acid glycosidase that catalyzes the removal of the terminal sialic acid residues of glycoconjugates (Verheijen et al 1982(Verheijen et al , 1985(Verheijen et al , 1987Hiraiwa et al 1988;van der Horst et al 1989). Mammalian lysosomal neuraminidase is associated with protective protein/cathepsin A (PPCA; EC 3.…”

Section: Introductionmentioning

confidence: 99%

Molecular and structural studies of Japanese patients with sialidosis type 1

et al. 2000

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“…Ganglioside GM1 13-galactosidase assays were carried out as described (20) using human liver 3-galactosidase purified to near homogeneity (21) as the enzyme source. A partially purified preparation of human placental sialidase prepared as described by Hiraiwa et al (22) was used as the source of sialidase. Sialidase assays were carried out as described (22) using gangliosides GM3 and GD1a as substrates, measuring released sialic acid by a sensitive fluorescence assay (23).…”

Section: Methodsmentioning

confidence: 99%

Binding and transport of gangliosides by prosaposin.

Hiraiwa

Soeda

Kishimoto

et al. 1992

Proc. Natl. Acad. Sci. U.S.A.

111

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“…The molecular mechanism of sialidase deficiency in galactosialidosis has never been characterized since until recently the lysosomal sialidase has not been cloned or purified. The sialidase activity has always been associated with the 1n27 MDa complex in the process of purification from tissues [16][17][18][19][20]. The dissociation of the complex resulted in the complete inactivation of sialidase.…”

Section: Introductionmentioning

confidence: 99%

Molecular mechanism of lysosomal sialidase deficiency in galactosialidosis involves its rapid degradation

Vinogradova

Michaud

Mezentsev

et al. 1998

Biochemical Journal

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Galactosialidosis is an inherited lysosomal storage disease caused by the combined deficiency of lysosomal sialidase and β-galactosidase secondary to the deficiency of cathepsin A/protective protein, which is associated with sialidase and β-galactosidase in a high-molecular weight (1.27 MDa) complex. Clinical phenotypes of patients as well as the composition of compounds which are stored in patient's tissues implicate sialidase deficiency as the underlying pathogenic defect. The recent cloning and sequencing of lysosomal sialidase [Pshezhetsky, Richard, Michaud, Igdoura, Wang, Elsliger, Qu, Leclerc, Gravel, Dallaire and Potier (1997), Nature Genet. 15, 316-320] allowed us to study the molecular mechanism of sialidase deficiency in galactosialidosis. By Western blotting, using antibodies against the recombinant human enzyme, and by NH2-terminal sequencing, we showed that sialidase is synthesized as a 45.5 kDa precursor and after the cleavage of the 47-amino acid signal peptide and glycosylation becomes a 48.3 kDa mature active enzyme present in the 1.27 kDa complex. Transgenic expression of sialidase in cultured skin fibroblasts from normal controls and from galactosialidosis patients, followed by immunofluorescent and immunoelectron microscopy showed that in both normal and affected cells the expressed sialidase was localized on lysosomal and plasma membranes, but the amount of sialidase found in galactosialidosis cells was ~ 5-fold reduced. Metabolic labelling studies demonstrated that the 48.3 kDa mature active form of sialidase was stable in normal fibroblasts (half-life ~ 2.7 h), whereas in galactosialidosis fibroblasts the enzyme was rapidly converted (half-life ~ 30 min) into 38.7 and 24 kDa catalytically inactive forms. Altogether our data provide evidence that the molecular mechanism of sialidase deficiency in galactosialidosis is associated with abnormal proteolytic cleavage and fast degradation.

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Human Placental Sialidase: Further Purification and Characterization

Cited by 40 publications

References 0 publications

Molecular and structural studies of Japanese patients with sialidosis type 1

Molecular and structural studies of Japanese patients with sialidosis type 1

Binding and transport of gangliosides by prosaposin.

Molecular mechanism of lysosomal sialidase deficiency in galactosialidosis involves its rapid degradation

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