Azido Groups Hamper Glycan Acceptance by Carbohydrate Processing Enzymes

Azido sugars have found frequent use as probes of biological systems in approaches ranging from cell surface metabolic labeling to activity-based proteomic profiling of glycosidases. However, little attention is typically paid to how well azide-substituted sugars represent the parent molecule, despite the substantial difference in size and structure of an azide compared to a hydroxyl. To quantitatively assess how well azides are accommodated, we have used glycosidases as tractable model enzyme systems reflecting what would also be expected for glycosyltransferases and other sugar binding/modifying proteins. In this vein, specificity constants have been measured for the hydrolysis of a series of azidodeoxy glucosides and N-acetylhexosaminides by a large number of glycosidases produced from expressed synthetic gene and metagenomic libraries. Azides at secondary carbons are not significantly accommodated, and thus, associated substrates are not processed, while those at primary carbons are productively recognized by only a small subset of the enzymes and often then only very poorly. Accordingly, in the absence of careful controls, results obtained with azide-modified sugars may not be representative of the situation with the natural sugar and should be interpreted with considerable caution. Azide incorporation can indeed provide a useful tool to monitor and detect glycosylation, but careful consideration should go into the selection of sites of azide substitution; such studies should not be used to quantitate glycosylation or to infer the absence of glycosylation activity.Bio-organic Synthesi

Lipid-mimicking phosphorus-based glycosidase inactivators as pharmacological chaperones for the treatment of Gaucher's disease

Gaucher's disease, the most prevalent lysosomal storage disorder, is caused by missense mutation of the GBA gene, ultimately resulting in deficient GCase activity, hence the excessive build-up of cellular glucosylceramide. Among different therapeutic strategies, pharmacological chaperoning of mutant GCase represents an attractive approach that relies on small organic molecules acting as protein stabilizers. Herein, we expand upon a new class of transient GCase inactivators based on a reactive 2-deoxy-2-fluoro-beta-d-glucoside tethered to an array of lipid-mimicking phosphorus-based aglycones, which not only improve the selectivity and inactivation efficiency, but also the stability of these compounds in aqueous media. This hypothesis was further validated with kinetic and cellular studies confirming restoration of catalytic activity in Gaucher cells after treatment with these pharmacological chaperones.Bio-organic Synthesi

Convergent synthesis of a fluorescence-quenched glycopeptide as a potential substrate for peptide: N-glycosidases

The fluorescence-quenched glycopeptide N-gamma-[2-acetamido-6-(2'-amino)benzamido-2,6-dideoxy-beta-D-glucopyranosyl]-N-alpha-benzoyl-Asn-Tyr(NO2)-OMe, having anthranilamide/3-nitrotyrosine as the donor/acceptor pair, was synthesized via a convergent route involving the EEDQ-mediated coupling of 2-acetamido-2,3-di-O-acetyl-6-(2'-tert-butoxycarbonylamino)benzamido-2,6-dideoxy-beta-D-glucopyranosylamine with N-benzoyl-Asp-Tyr(NO2)-OMe, followed by deprotection. (C) 1998 Elsevier Science Ltd. All rights reserved

Mechanistic analysis of the blood group antigen-cleaving endo-β-galactosidase from <i>Clostridium perfringens</i>

International audienc

Mechanistic analysis of the blood group antigen-cleaving endo-β-galactosidase from <i>Clostridium perfringens</i>

International audienc

N-Glycosyl phosphonamidates: potential transition-state analogue inhibitors of glycopeptidases

The synthesis of N-glycosyl phosphonamidates has been accomplished via the coupling of peracetylated glycosylamines with an appropriate phosphonochloridate in the presence of pyridine. The resulting glycosyl phosphonamidate esters are dealkylated with bromotrimethylsilane and then deacetylated to give the target compounds, which are potential transition-state analogue inhibitors of glycopeptidases and may prove useful as haptens for generating catalytic antibodies with glycopeptidase activity