The use of glycosides of 6- and 8-acylamino-4-methylumbelliferone in studies of the specificity and properties of human lysosomal glycolipid hydrolases

Wiederschain, G.Ya.; Kozlova, I. K.; Ільїна, Галина Самуіловна; Mikhaylova, Margarita A.; Beyer, E.M.

doi:10.1016/0008-6215(92)84111-5

Cited by 12 publications

(7 citation statements)

References 33 publications

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“…The results were consistent with the range of values previously reported in the literature (17,26,30 ) or routinely generated through conventional methods in laboratory facilities specializing in prenatal diagnostics (G. Gaslini Institute, Diagnosi PrePostnatale Malattie Metaboliche Laboratori, Genova, Italy (31 )) ( Table 1). In particular, GALC specific activity was 4.32 (0.41) mU/mg and 0.14 (0.025) mU/mg in healthy controls and Krabbe fibroblasts, respectively.…”

Section: Galc Assay Applicationssupporting

confidence: 91%

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Specific Determination of β-Galactocerebrosidase Activity via Competitive Inhibition of β-Galactosidase

et al. 2009

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BACKGROUND:The determination of cellular ␤-galactocerebrosidase activity is an established procedure to diagnose Krabbe disease and monitor the efficacy of gene/stem cell-based therapeutic approaches aimed at restoring defective enzymatic activity in patients or disease models. Current biochemical assays for ␤-galactocerebrosidase show high specificity but generally require large protein amounts from scanty sources such as hematopoietic or neural stem cells. We developed a novel assay based on the hypothesis that specific measurements of ␤-galactocerebrosidase activity can be performed following complete inhibition of ␤-galactosidase activity.

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Section: Galc Assay Applicationssupporting

confidence: 91%

“…We separated ␤-Gal from GALC using gel filtration chromatography (26,27 ). To this end, wt or twi brain tissue extracts were loaded onto a Sephadex S-300 gel filtration column (Fig.…”

Section: Partial Purification Of ␤-Galactocerebrosidasementioning

confidence: 99%

Specific Determination of β-Galactocerebrosidase Activity via Competitive Inhibition of β-Galactosidase

et al. 2009

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“…To examine the GALC enzyme kinetics in the presence of the small‐molecule candidates, human GALC was purified from BHK‐21, as described in Materials and Methods. Because the HMU substrate demonstrated considerable variability resulting from a prolonged hydrolysis of the HMU synthetic substrate (Wiederschain et al, ), 4MUβGal, another fluorogenic synthetic substrate, was utilized for the kinetic examinations of GALC (Supp. Info.…”

Section: Resultsmentioning

confidence: 99%

Cell‐based high‐throughput screening identifies galactocerebrosidase enhancers as potential small‐molecule therapies for Krabbe's disease

Jang

Tian

et al. 2016

J of Neuroscience Research

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Krabbe disease, also known as globoid-cell leukodystrophy (GLD), is a lysosomal storage disease (LSD) caused by the deficiency of the lysosomal enzyme β-galactocerebrosidase (GALC), resulting in severe neurological manifestations related to demyelination secondary to elevated galactosylsphingosine (psychosine) with its subsequent cytotoxicity. The only available treatment is hematopoietic stem cell transplantation that delays the disease onset but has not prevented further long-term neurological manifestations. Here, we report the identification of small molecules that enhance mutant GALC activity identified by quantitative cell-based high-throughput screen (qHTS). Using a specific neurologically relevant murine cell-line (145M-Twi) modified to express common human hGALC-G270D mutant, we were able to detect GALC activity in 1,536-well microplate format. The qHTS of approximately 46,000 compounds identified 3 small molecules that show significant enhancements of residual mutant GALC activity in primary cell lines from GLD patients. Interestingly, these compounds showed to increase the levels of GALC-G270D mutant in the lysosomal compartment. In kinetic assessments, these small molecules fail to disturb GALC kinetics profile at acidic conditions, which is highly desirable for folding-assisting molecules operating in the endoplasmic reticulum and not affecting GALC catalytic properties once in the lysosomal compartment. In addition, these small molecules rescue the decreased GALC activity at neutral pH and partially stabilize GALC at heat-denaturating conditions. These are drug-like compounds that can be used as the starting point to develop novel small molecule agents to treat the progressive neurodegenerative course of GLD.

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“…4-Methylumbelliferyl aglycone analogues were also explored to better suit the properties of glycosidases [71][72][73][74]. A number of other fluorenegic glycosides based on aglycons other than 4-methylumbelliferyl were shown to be useful glycosidase substrates, possessing properties that could be more suitable for their respective applications [75][76][77].…”

Section: Fluorogenic Glycosidase Substratesmentioning

confidence: 98%

Fluorescently labelled glycans and their applications

Yan

Yalagala

Yan³

2015

Glycoconj J

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This review summarises the literature on the synthesis and applications of fluorescently labelled carbohydrates. Due to the sensitivity of fluorescent detection, this approach provides a useful tool to study processes involving glycans. A few general categories of labelling are presented, in situ labelling of carbohydrates with fluorophores, fluorescently labelled glycolipids, fluorogenic glycans, pre-formed fluorescent glycans for intracellular applications, glycan-decorated fluorescent polymers, fluorescent glyconanoparticles, and other functional fluorescent glycans.

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The use of glycosides of 6- and 8-acylamino-4-methylumbelliferone in studies of the specificity and properties of human lysosomal glycolipid hydrolases

Cited by 12 publications

References 33 publications

Specific Determination of β-Galactocerebrosidase Activity via Competitive Inhibition of β-Galactosidase

Specific Determination of β-Galactocerebrosidase Activity via Competitive Inhibition of β-Galactosidase

Cell‐based high‐throughput screening identifies galactocerebrosidase enhancers as potential small‐molecule therapies for Krabbe's disease

Fluorescently labelled glycans and their applications

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