N-Azidoacetylmannosamine-mediated chemical tagging of gangliosides

Bussink, Anton P.; Swieten, Paul F. van; Ghauharali, Karen; Scheij, Saskia; Eijk, Marco van; Wennekes, Tom; Marel, Gijs A. van der; Boot, Rolf G.; Aerts, Johannes M. F. G.; Overkleeft, Herman S.

doi:10.1194/jlr.c700006-jlr200

Cited by 24 publications

(29 citation statements)

References 23 publications

(16 reference statements)

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“…We next tested if the new photocrosslinking sialic acids could also be incorporated into sialic acid-containing glycolipids, as has been shown for other sialic acid analogs. 13, 36–41 Briefly, Jurkat cells were cultured in the presence of sugars 1–6 for 72 hours. After harvesting the cells, a series of extractions were employed to remove insoluble membrane components, proteins, phospholipids, and uncharged lipids.…”

Section: Resultsmentioning

confidence: 99%

Metabolism of Diazirine-Modified N-Acetylmannosamine Analogues to Photo-Cross-Linking Sialosides

et al. 2011

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Terminal sialic acid residues often mediate the interactions of cell surface glycoconjugates. Sialic acid-dependent interactions typically exhibit rapid dissociation rates, precluding the use of traditional biological techniques for complex isolation. To stabilize these transient interactions, we employ a targeted photocrosslinking approach in which a diazirine photocrosslinker is incorporated into cell surface sialylated glycoconjugates through the use of metabolic oligosaccharide engineering. We describe three diazirine-modified N-acetylmannosamine (ManNAc) analogs in which the length of the linker between the pyranose ring and the diazirine was varied. These analogs were each metabolized to their respective sialic acid counterparts, which were added to both glycoproteins and glycolipids. Diazirine-modified sialic acid analogs could be incorporated into both α2–3 and α2–6 linkages. Upon exposure to UV irradiation, diazirine-modified glycoconjugates were covalently crosslinked to their interaction partners. We demonstrate that all three diazirine-modified analogs were capable of competing with endogeneous sialic acid, albeit to varying degrees. We found that larger analogs were less efficiently metabolized, yet could still function as effective crosslinkers. Notably, the addition of the diazirine substituent interferes with metabolism of ManNAc analogs to glycans other than sialosides, providing fidelity to selectively incorporate the crosslinker into sialylated molecules. These compounds are non-toxic and display only minimal growth inhibition at the concentrations required for crosslinking studies. This report provides essential information for the deployment of photocrosslinking analogs to capture and study ephemeral, yet essential, sialic acid-mediated interactions.

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Section: Resultsmentioning

confidence: 99%

Metabolism of Diazirine-Modified N-Acetylmannosamine Analogues to Photo-Cross-Linking Sialosides

et al. 2011

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“…For example, Wong and coworkers [61] labeled cells with fucose analogues containing either an azide or an alkyne functionality and then imaged the cells with fluorogenic alkyne or azide reagents, respectively, using CuAAC [57,61]. Overkleeft and coworkers recently demonstrated that a significant portion of the labeled sialic acids in Jurkat cells treated with ManNAz reside within glycolipids [65], suggesting the possibility of imaging their dynamics using bioorthogonal click chemistries as well.…”

Section: Metabolic Labeling Of Glycans and Other Posttranslational Momentioning

confidence: 97%

Bioorthogonal Click Chemistry: Covalent Labeling in Living Systems

2007

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The term "click chemistry" defines a powerful set of chemical reactions that are rapid, selective, and high-yielding. These reactions, some of which are less than 10 years old, have been applied in diverse areas, including drug discovery, materials science, and chemical biology. In chemical biology, click chemistry has been used in the selective labeling of biomolecules within living systems, allowing proteins, glycans, and other important biomolecules to be monitored in a physiologically relevant environment rather than in an in vitro setting. This demanding application requires not only the aforementioned characteristics of click chemistry but, additionally, that the reactions are bioorthogonal -that is, non-interacting with biological functionality while proceeding under physiological conditions -and that the reagents are non-toxic. Of the many extant click reactions, only a select few possess this unique combination of attributes, notably the Staudinger ligation of azides and triarylphosphines and [3 þ 2] dipolar cycloadditions of azides with strained alkynes. This minireview describes the characteristics of bioorthogonal click reactions as well as recent applications toward labeling biomolecules in cells and living organisms.

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“…Succinimidyl hept-6-ynoate (8) Synthesis of Azidoglucosylceramide 2. 2,3,4-Tri-Oacetyl-6-azido-6-deoxy-D-glucopyranose (32). Glucose derivative 31 48 was subjected to anomeric deprotection following the procedure described by Zhang and Kovać.…”

Section: ■ Conclusionmentioning

confidence: 99%

Synthetic Glycosphingolipids for Live-Cell Labeling

et al. 2016

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Glycosphingolipids are an important component of cell membranes that are involved in many biological processes. Fluorescently labeled glycosphingolipids are frequently used to gain insight into their localization. However, the attachment of a fluorophore to the glycan part or-more commonly-to the lipid part of glycosphingolipids is known to alter the biophysical properties and can perturb the biological function of the probe. Presented here is the synthesis of novel glycosphingolipid probes with mono- and disaccharide head groups and ceramide moieties containing fatty acids of varying chain length (C4 to C20). These glycosphingolipids bear an azide or an alkyne group as chemical reporter to which a fluorophore can be attached through a bioorthogonal ligation reaction. The fluorescent tag and any linker connected to it can be chosen in a flexible manner. We demonstrate the suitability of the probes by selective visualization of the plasma membrane of living cells by confocal microscopy techniques. Whereas the derivatives with the shorter fatty acids can be directly applied to HEK 293T cells, the hydrophobic glycosphingolipids with longer fatty acids can be delivered to cells using fusogenic liposomes.

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N-Azidoacetylmannosamine-mediated chemical tagging of gangliosides

Cited by 24 publications

References 23 publications

Metabolism of Diazirine-Modified N-Acetylmannosamine Analogues to Photo-Cross-Linking Sialosides

Metabolism of Diazirine-Modified N-Acetylmannosamine Analogues to Photo-Cross-Linking Sialosides

Bioorthogonal Click Chemistry: Covalent Labeling in Living Systems

Synthetic Glycosphingolipids for Live-Cell Labeling

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