A Time-Resolved Immunofluorometric Assay of Sialyl Lewis x-Degrading α2,3-Sialidase Activity

Räbinä, Jarkko; Pikkarainen, Mari; Miyasaka, Masayuki; Renkonen, Risto

doi:10.1006/abio.1998.2590

Cited by 10 publications

(4 citation statements)

References 34 publications

(27 reference statements)

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“…Currently the cellular localization and the type of endothelial sialidase(s) are not known. However, we have shown that the endothelial sialidase activity has a pH optimum around 4.0 suggesting that it is of lysosomal type ([22] and data not shown) and that desialylation most likely occurs in lysosomes after internalization from the cell surface.…”

Section: Resultsmentioning

confidence: 84%

High endothelial cells synthesize and degrade sLex. Putative implications for L‐selectin‐dependent recognition

et al. 1999

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L-selectin guides lymphocytes into peripheral lymphoid tissues by recognizing glycoprotein ligands decorated with 6-sulfated sialyl Lewis x (sulfo sLex). Here we have used a rat peripheral lymph node high endothelial cell line (Ax) to study in detail the synthesis, expression and degradation of sLex epitope. We show here that Ax cells possess active K K(1,3)fucosyltransferase Fuc-TVII, the enzyme responsible for the final fucosylation of sialyl-N-acetyllactosamine during sLex synthesis, and express sLex on the cell surface. Furthermore, these cells degrade sLex, primarily by desialylating it to neutral Lex epitopes by K K(2,3)sialidase(s).z 1999 Federation of European Biochemical Societies.

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Section: Resultsmentioning

confidence: 84%

High endothelial cells synthesize and degrade sLex. Putative implications for L‐selectin‐dependent recognition

et al. 1999

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“…We have earlier used similar solution-phase assay format for measuring sialidase activity (17). In the FucT assay that we earlier published, we used immobilized acceptor (16), but later we have noticed that soluble acceptor is clearly better for this enzyme (not shown).…”

Section: Discussionmentioning

confidence: 96%

“…We have recently developed rapid and sensitive high-throughput assays for glycosyltransferases and glycosidases, which employ microplate assay technology and time-resolved fluorometric detection (16,17). In the detection of GDP-Fuc, we utilized ␣1,3-fucosyltransferase assay, in which the limiting factor was the presence or absence of GDP-Fuc in the sample.…”

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confidence: 99%

Two Time-Resolved Fluorometric High-Throughput Assays for Quantitation of GDP--Fucose

Räbinä

Mattila

Renkonen

2000

Analytical Biochemistry

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“…Hyposialylation of vWF in pulmonary hypertension could be due to abnormal intracellular or extracellular processing or both. Mammalian sialidases have been identified in several cell types including endothelial cells (23) and have been shown to be involved in modulating a broad range of cellular processes. Although most sialidases are cytosolic or lysosomal (24), sialidase activities have been identified in the supernatant of cultured cells (25).…”

Section: Discussionmentioning

confidence: 99%

Decreased Sialic Acid Content of Plasma von Willebrand Factor in Precapillary Pulmonary Hypertension

B²,

Ny³

2000

Thromb Haemost

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SummaryIn pulmonary hypertension, defective von Willebrand factor protein (vWF) lacking large multimers is present in circulation. This is associated with evidence of chronic endogenous platelet activation. Since asialo vWF has been shown to promote platelet activation and aggregation, we decided to investigate possible changes in the sialic acid content of plasma vWF in patients with precapillary pulmonary hypertension. vWF-associated sialic acid was measured indirectly as a wheat germ agglutinin-reactive substance (WGA-RS, Western blotting), and directly, as a thiobarbituric acid-reactive substance (TBA-RS, spectrophotometric reading). In the sixteen patients we studied (ages 8–45 yr), circulating vWF concentration was 2.18 times normal (p <0.001). However, patient vWF subunit contained 19% (WGA-RS) to 24% (TBA-RS) less sialic acid than the normal protein (p <0.05 for both determinations). In five patients, vWF-associated sialic acid was below 50% normal. We conclude that circulating vWF is hyposialylated in precapillary pulmonary hypertension and speculate that this might influence its interaction with platelets in vivo in these patients.

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A Time-Resolved Immunofluorometric Assay of Sialyl Lewis x-Degrading α2,3-Sialidase Activity

Cited by 10 publications

References 34 publications

High endothelial cells synthesize and degrade sLex. Putative implications for L‐selectin‐dependent recognition

High endothelial cells synthesize and degrade sLex. Putative implications for L‐selectin‐dependent recognition

Two Time-Resolved Fluorometric High-Throughput Assays for Quantitation of GDP--Fucose

Decreased Sialic Acid Content of Plasma von Willebrand Factor in Precapillary Pulmonary Hypertension

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