1361993-37-4Relevant articles and documents
Metabolic glycoengineering in hmsc-tert as a model for skeletal precursors by using modified azide/alkyne monosaccharides
Altmann, Stephan,Mut, Jürgen,Wolf, Natalia,Mei?ner-Weigl, Jutta,Rudert, Maximilian,Jakob, Franz,Gutmann, Marcus,Lühmann, Tessa,Seibel, Jürgen,Ebert, Regina
, p. 1 - 16 (2021)
Metabolic glycoengineering enables a directed modification of cell surfaces by introducing target molecules to surface proteins displaying new features. Biochemical pathways involving glycans differ in dependence on the cell type; therefore, this technique should be tailored for the best results. We characterized metabolic glycoengineering in telomerase-immortalized human mesen-chymal stromal cells (hMSC-TERT) as a model for primary hMSC, to investigate its applicability in TERT-modified cell lines. The metabolic incorporation of N-azidoacetylmannosamine (Ac4Man-NAz) and N-alkyneacetylmannosamine (Ac4ManNAl) into the glycocalyx as a first step in the gly-coengineering process revealed no adverse effects on cell viability or gene expression, and the in vitro multipotency (osteogenic and adipogenic differentiation potential) was maintained under these adapted culture conditions. In the second step, glycoengineered cells were modified with flu-orescent dyes using Cu-mediated click chemistry. In these analyses, the two mannose derivatives showed superior incorporation efficiencies compared to glucose and galactose isomers. In time-de-pendent experiments, the incorporation of Ac4ManNAz was detectable for up to six days while Ac4ManNAl-derived metabolites were absent after two days. Taken together, these findings demonstrate the successful metabolic glycoengineering of immortalized hMSC resulting in transi-ent cell surface modifications, and thus present a useful model to address different scientific ques-tions regarding glycosylation processes in skeletal precursors.
Diazo groups endure metabolism and enable chemoselectivity in cellulo
Andersen, Kristen A.,Aronoff, Matthew R.,McGrath, Nicholas A.,Raines, Ronald T.
, p. 2412 - 2415 (2015)
We introduce a stabilized diazo group as a reporter for chemical biology. ManDiaz, which is a diazo derivative of N-acetylmannosamine, is found to endure cellular metabolism and label the surface of a mammalian cell. There its diazo group can undergo a 1,3-dipolar cycloaddition with a strained alkyne, providing a signal comparable to that from the azido congener, ManNAz. The chemoselectivity of diazo and alkynyl groups enables dual labeling of cells that is not possible with azido and alkynyl groups. Thus, the diazo group, which is approximately half the size of an azido group, provides unique opportunities for orthogonal labeling of cellular components.
Design, synthesis and cytotoxic activity of N-Modified oleanolic saponins bearing A glucosamine
Lin, You-Yu,Chan, She-Hung,Juang, Yu-Pu,Hsiao, Hsin-Min,Guh, Jih-Hwa,Liang, Pi-Hui
, p. 1942 - 1958 (2017/11/16)
A series of N-acyl, N-alkoxycarbonyl, and N-alkylcarbamoyl derivatives of 2′-deoxy-glucosyl bearing oleanolic saponins were synthesized and evaluated against HL-60, PC-3, and HT29 tumor cancer cells. The SAR studies revealed that the activity increased in order of conjugation of 2′ -amino group with carbamate > amide > urea derivatives. Lengthening the alkyl chain increased the cytotoxicity, the peak activity was found to around heptyl to nonyl substitutions. 2′-N-heptoxycarbonyl derivative 56 was found to be the most cytotoxic (IC50 = 0.76 μM) against HL-60 cells. Due to the interesting SARs of alkyl substitutions, we hypothesized that their location in the cell was different, and pursued a location study using 2′-(4″-pentynoylamino) 2′-deoxy-glucosyl OA, which suggested that these compounds distributed mainly in the cytosol.
Alkynyl sugar analogs for labeling and visualization of glycoconjugates in cells
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Page/Page column 23; 32, (2017/12/09)
Methods for metabolic oligosaccharide engineering that incorporates derivatized alkyne-bearing sugar analogs as “tags” into cellular glycoconjugates are disclosed. Alkynyl derivatized Fuc and alkynyl derivatized ManNAc sugars are incorporated into cellular glycoconjugates. Chemical probes comprising an azide group and a visual or fluorogenic probe and used to label alkyne-derivatized sugar-tagged glycoconjugates are disclosed. Chemical probes bind covalently to the alkynyl group by Cu(I)-catalyzed [3+2] azide-alkyne cycloaddition and are visualized at the cell surface, intracellularly, or in a cellular extract. The labeled glycoconjugate is capable of detection by flow cytometry, SDS-PAGE, Western blot, ELISA, confocal microscopy, and mass spectrometry.
Chemical probes for the functionalization of polyketide intermediates
Riva, Elena,Wilkening, Ina,Gazzola, Silvia,Li, W. M. Ariel,Smith, Luke,Leadlay, Peter F.,Tosin, Manuela
supporting information, p. 11944 - 11949 (2015/02/19)
A library of functionalized chemical probes capable of reacting with ketosynthase-bound biosynthetic intermediates was prepared and utilized to explore in vivo polyketide diversification. Fermentation of ACP mutants of S. lasaliensis in the presence of the probes generated a range of unnatural polyketide derivatives, including novel putative lasalocid A derivatives characterized by variable aryl ketone moieties and linear polyketide chains (bearing alkyne/azide handles and fluorine) flanking the polyether scaffold. By providing direct information on microorganism tolerance and enzyme processing of unnatural malonyl-ACP analogues, as well as on the amenability of unnatural polyketides to further structural modifications, the chemical probes constitute invaluable tools for the development of novel mutasynthesis and synthetic biology.
Cellular consequences of copper complexes used to catalyze bioorthogonal click reactions
Kennedy, David C.,McKay, Craig S.,Legault, Marc C. B.,Danielson, Dana C.,Blake, Jessie A.,Pegoraro, Adrian F.,Stolow, Albert,Mester, Zoltan,Pezacki, John Paul
, p. 17993 - 18001 (2011/12/16)
Copper toxicity is a critical issue in the development of copper-based catalysts for copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) reactions for applications in living systems. The effects and related toxicity of copper on mammalian cells are dependent on the ligand environment. Copper complexes can be highly toxic, can induce changes in cellular metabolism, and can be rapidly taken up by cells, all of which can affect their ability to function as catalysts for CuAAC in living systems. Herein, we have evaluated the effects of a number of copper complexes that are typically used to catalyze CuAAC reactions on four human cell lines by measuring mitochondrial activity based on the metabolism of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) to study toxicity, inductively coupled plasma mass spectrometry to study cellular uptake, and coherent anti-Stokes Raman scattering (CARS) microscopy to study effects on lipid metabolism. We find that ligand environment around copper influences all three parameters. Interestingly, for the Cu(II)-bis-l-histidine complex (Cu(his)2), cellular uptake and metabolic changes are observed with no toxicity after 72 h at micromolar concentrations. Furthermore, we show that under conditions where other copper complexes kill human hepatoma cells, Cu(I)-l-histidine is an effective catalyst for CuAAC labeling of live cells following metabolic incorporation of an alkyne-labeled sugar (Ac 4ManNAl) into glycosylated proteins expressed on the cell surface. This result suggests that Cu(his)2 or derivatives thereof have potential for in vivo applications where toxicity as well as catalytic activity are critical factors for successful bioconjugation reactions.
