Stabilization of Glucocerebrosidase by Active Site Occupancy

Bdira, Fredj Ben; Kallemeijn, Wouter W.; Oussoren, Saskia V.; Scheij, Saskia; Bleijlevens, Boris; Florea, Bogdan I.; Roomen, Cindy P. A. A. van; Ottenhoff, Roelof; Kooten, Mariëlle van; Walvoort, Marthe T. C.; Witte, Martin D.; Boot, Rolf G.; Ubbink, Marcellus; Overkleeft, Herman S.; Aerts, Johannes M. F. G.

doi:10.1021/acschembio.7b00276

Cited by 25 publications

(21 citation statements)

References 58 publications

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“…These results were confirmed using a recently developed ultrasensitive fluorescent probe, MDW941, which specifically reacts with active forms of the enzyme and has been shown to be sensitive enough to detect the activity of recombinant GBA in the attomolar range (48)(49)(50). Using this probe, an even greater disparity between GBA activity in WT and Grn -/tissues was observed ( Fig.…”

Section: Pgrn Deficiency Leads To Reduced Gba Activitiesmentioning

confidence: 62%

Progranulin deficiency leads to reduced glucocerebrosidase activity

Zhou

Paushter

Pagan

et al. 2019

Preprint

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Mutation in the GRN gene, encoding the progranulin (PGRN) protein, shows a dose-dependent disease correlation, wherein haploinsufficiency results in frontotemporal lobar degeneration (FTLD) and complete loss results in neuronal ceroid lipofuscinosis (NCL). Although the exact function of PGRN is unknown, it has been increasingly implicated in lysosomal physiology. Here we report that PGRN interacts with the lysosomal enzyme, glucocerebrosidase (GBA), and is essential for proper GBA activity. GBA activity is significantly reduced in tissue lysates from PGRN-deficient mice. This is further evidence that reduced lysosomal hydrolase activity may be a pathological mechanism in cases of GRN-related FTLD and NCL.

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Section: Pgrn Deficiency Leads To Reduced Gba Activitiesmentioning

confidence: 62%

Progranulin deficiency leads to reduced glucocerebrosidase activity

Zhou

Paushter

Pagan

et al. 2019

Preprint

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“…The IFG rigidification effects on GBA1 seem to propagate beyond its binding pocket to surrounding regions, thus restricting the local protein dynamics. Recently, the occupancy effect of the binding site in GBA1 by small hydrophilic and amphiphilic mechanism-based inhibitors was studied (88). The complex of GBA1 with the small glycomimetic conduritol B epoxide (CBE) was found to cause only minor changes in the enzyme fluorescence spectrum.…”

Section: Therapeutic Noncovalent Inhibitorsmentioning

confidence: 99%

“…Residues residing in the core of the TIM barrel domain were highly protected against the HDX, and loops connecting the -helices and -sheet of the catalytic domain showed the lowest protection (87). The conformational stability and rigidity of GBA1 is pHdependent, as indicated by measuring the melting temperature (T m ) at different pH values and resistance toward tryptic digestion (88). At acidic pH, similar to that of lysosomes, the T m of GBA1 is 4°C higher than at neutral pH, as found in the ER.…”

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confidence: 99%

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Distinguishing the differences in β-glycosylceramidase folds, dynamics, and actions informs therapeutic uses

Bdira

Artola

Overkleeft

et al. 2018

Journal of Lipid Research

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Glycoconjugates play essential roles in diverse biological processes and abnormalities and have been linked to multiple pathologies. The staggering structural diversity of glycoconjugates stems from the almost infinite possible combinations by which multiple monosaccharide building blocks may be linked with each other (1). For instance, 1,056 unique trisaccharides can be formed just from 3 different monosaccharides. Polysaccharides are very stable compounds: their spontaneous hydrolysis takes place at a rate of 10 15 s 1 , corresponding to a half-life of 4.7 million years (2). To allow for the efficient metabolism of glycoconjugates, enzymes have evolved as specialized catalysts. In most organisms, an estimated 1% to 3% of the genes encode carbohydrate-active enzymes (3). Among these are glycoside hydrolases (GHs) that can enhance the rate of hydrolysis of specific carbohydrate glycosidic bonds in glycoconjugates more than 10 17 times (2). These GH enzymes show marked specificity regarding number, position, and configuration of the hydroxyl groups in their substrate sugar. They are widely applied in biotechnology, for example, in biofuel production, paper pulp bleaching, and the food industry (4, 5). Likewise, specific GH inhibitors are extensively used as agrochemicals and therapeutic agents (6-8). Abnormalities in GHs underlie metabolic disorders in humans, for instance, inherited lysosomal storage disorders and lactose intolerance (9, 10). GH enzymes differ in substrate specificity, mode of enzymatic attack Abstract Glycosyl hydrolases (GHs) are carbohydrate-active enzymes that hydrolyze a specific -glycosidic bond in glycoconjugate substrates; -glucosidases degrade glucosylceramide, a ubiquitous glycosphingolipid. GHs are grouped into structurally similar families that themselves can be grouped into clans. GH1, GH5, and GH30 glycosidases belong to clan A hydrolases with a catalytic (/) 8 TIM barrel domain, whereas GH116 belongs to clan O with a catalytic (/) 6 domain. In humans, GH abnormalities underlie metabolic diseases. The lysosomal enzyme glucocerebrosidase (family GH30), deficient in Gaucher disease and implicated in Parkinson disease etiology, and the cytosol-facing membrane-bound glucosylceramidase (family GH116) remove the terminal glucose from the ceramide lipid moiety. Here, we compare enzyme differences in fold, action, dynamics, and catalytic domain stabilization by binding site occupancy. We also explore other glycosidases with reported glycosylceramidase activity, including human cytosolic -glucosidase, intestinal lactase-phlorizin hydrolase, and lysosomal galactosylceramidase. Last, we describe the successful translation of research to practice: recombinant glycosidases and glucosylceramide metabolism modulators are approved drug products (enzyme replacement therapies). Activity-based probes now facilitate the diagnosis of enzyme deficiency and screening for compounds that interact with the catalytic pocket of glycosidases. Future research may deepen the understanding of the function...

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“…In one study, the effect of miglustat was attributed to the rescue of the delF508‐CTRF protein by inhibiting its deglucosylation (inhibition of ER α‐1,2‐glucosidase) and the subsequent del508‐CFTR/calnexin interaction, a process that leads to protein degradation . In other structure–activity studies, hydrophobic groups were tethered to six‐ and seven‐membered DNJ analogues, which functioned as pharmacological chaperones to improve folding of mutant GBA1 in Gaucher's disease and to promote GBA1 trafficking from the endoplasmic reticulum to the lysosome …”

Section: Introductionmentioning

confidence: 99%

Structure–Activity Studies of N‐Butyl‐1‐deoxynojirimycin (NB‐DNJ) Analogues: Discovery of Potent and Selective Aminocyclopentitol Inhibitors of GBA1 and GBA2

Gupta

Yang

et al. 2017

ChemMedChem

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Analogues of N‐butyl‐1‐deoxynojirimycin (NB‐DNJ) were prepared and assayed for inhibition of ceramide‐specific glucosyltransferase (CGT), non‐lysosomal β‐glucosidase 2 (GBA2) and the lysosomal β‐glucosidase 1 (GBA1). Compounds 5 a–6 f, which carry sterically demanding nitrogen substituents, and compound 13, devoid of the C3 and C5 hydroxy groups present in DNJ/NB‐DGJ (N‐butyldeoxygalactojirimycin) showed no inhibitory activity for CGT or GBA2. Inversion of stereochemistry at C4 of N‐(n‐butyl)‐ and N‐(n‐nonyl)‐DGJ (compounds 24) also led to a loss of activity in these assays. The aminocyclopentitols N‐(n‐butyl)‐ (35 a), N‐(n‐nonyl)‐4‐amino‐5‐(hydroxymethyl)cyclopentane‐ (35 b), and N‐(1‐(pentyloxy)methyl)adamantan‐1‐yl)‐1,2,3‐triol (35 f), were found to be selective inhibitors of GBA1 and GBA2 that did not inhibit CGT (>1 mm), with the exception of 35 f, which inhibited CGT with an IC50 value of 1 mm. The N‐butyl analogue 35 a was 100‐fold selective for inhibiting GBA1 over GBA2 (K i values of 32 nm and 3.3 μm for GBA1 and GBA2, respectively). The N‐nonyl analogue 35 b displayed a K i value of ≪14 nm for GBA1 inhibition and a K i of 43 nm for GBA2. The N‐(1‐(pentyloxy)methyl)adamantan‐1‐yl) derivative 35 f had K i values of ≈16 and 14 nm for GBA1 and GBA2, respectively. The related N‐bis‐substituted aminocyclopentitols were found to be significantly less potent inhibitors than their mono‐substituted analogues. The aminocyclopentitol scaffold should hold promise for further inhibitor development.

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Stabilization of Glucocerebrosidase by Active Site Occupancy

Cited by 25 publications

References 58 publications

Progranulin deficiency leads to reduced glucocerebrosidase activity

Progranulin deficiency leads to reduced glucocerebrosidase activity

Distinguishing the differences in β-glycosylceramidase folds, dynamics, and actions informs therapeutic uses

Structure–Activity Studies of N‐Butyl‐1‐deoxynojirimycin (NB‐DNJ) Analogues: Discovery of Potent and Selective Aminocyclopentitol Inhibitors of GBA1 and GBA2

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