Irradiation of cells with UV light activated the crosslinker, resulting in formation of covalent bonds between O-GlcNAc-modified proteins and neighboring molecules, which could be identified by mass spectrometry. A preliminary study of the mechanism of this reaction, and refined conditions for its in vivo execution, are reported. Aldehyde- and ketone-functionalized proteins are appealing substrates for the development of chemically modified biotherapeutics and protein-based materials. Fluorescent probes designed for activation by bioorthogonal chemistry have enabled the visualization of biomolecules in living systems. Herein, we designed and synthesized the first chemically defined ligands for dectin-1 and dectin-2. Using an in vitro biochemical assay, we confirmed that Rv3406 is a sulfatase with a preference for alkyl sulfate substrates similar to those processed by AtsK. View details for DOI 10.1126/science.1155106, View details for Web of Science ID 000255454300046. The ESI-MS assay is used to determine NodST kinetic parameters, including K(M), V(max), and K(i) (for PAP). A., Riley, L. W., Bertozzi, C. R. 5 '-Adenosinephosphosulphate reductase (CysH) protects Mycobacterium tuberculosis against free radicals during chronic infection phase in mice. The alkyl azide was then selectively derivatized with cyclo-octyne conjugates to various probes. Riley, N. M., Malaker, S. A., Bertozzi, C. R. Sensitivity optimisation of tuberculosis bioaerosol sampling. Among the EMBs loci were genes encoding RipC and the FtsEX complex, a PG cleaving enzyme required for proper cell division and its predicted regulator, respectively. The route developed for the model compound can be readily extended to the synthesis of native SL-I as well as additional analogues for use in the investigation of SL-I's functions. The Staudinger ligation of azides and phosphines has found widespread use in the field of chemical biology, but the mechanism of the transformation has not been characterized in detail. View details for DOI 10.1074/jbc.M405709200, View details for Web of Science ID 000223791500101, View details for Web of Science ID 000225338800003. Zebrafish embryos were treated with an unnatural sugar to metabolically label their cell-surface glycans with azides. A key feature of SiaLev is the presence of a ketone group that can be exploited in chemoselective ligation reactions to deliver small-molecule probes to the cell surface. View details for DOI 10.1016/j.biomaterials.2004.01.025, View details for Web of Science ID 000222040300008, View details for DOI 10.1002/cbic.200400156, View details for Web of Science ID 000224453500014. We provide evidence that conserved switch motifs in the G domain of CysN allosterically mediate interactions between the nucleotide binding sites. To quickly assess the protein engineering outcomes, we developed a new platform for quantifying extracellular alkyne-tagged metabolites through a fluorogenic click reaction. View details for DOI 10.1073/pnas.2335201100, View details for Web of Science ID 000187227200045, View details for PubMedCentralID PMC299823. View details for Web of Science ID 000430563200441. Here, we describe a method for visualizing and analyzing organelle- and stimulus-specific O-GlcNAcylated proteins and use it to identify the mitochondrial voltage-dependent anion channel 2 (VDAC2) as an O-GlcNAc substrate. View details for Web of Science ID 000250428200015. In the ligation reaction, the intermediate aza-ylide undergoes intramolecular reaction with an ester, forming an amide bond faster than aza-ylide hydrolysis would otherwise occur in water. Synthetic mimics of the complex assemblies found on cell surfaces can modulate cellular interactions and are under development as therapeutic agents. Converse, S. E., Mougous, J. D., Leavell, M. D., Leary, J. Molecular imaging enables visualization of specific molecules in vivo and without substantial perturbation to the target molecule's environment. Siegrist, M. S., Whiteside, S., Jewett, J. C., Aditham, A., Cava, F., Bertozzi, C. R. Strategy for Dual-Analyte Luciferin Imaging: In Vivo Bioluminescence Detection of Hydrogen Peroxide and Caspase Activity in a Murine Model of Acute Inflammation. We describe the application of click-EM in imaging metabolically tagged DNA, RNA and lipids in cultured cells and neurons and highlight its use in tracking peptidoglycan synthesis in the Gram-positive bacterium Listeria monocytogenes. Likewise, these crosslinking probes serve as ideal chemical tools for structural studies between NRPS modules where functional assays are lacking. View details for DOI 10.1002/pmic.200900708, View details for Web of Science ID 000273515000001, View details for PubMedCentralID PMC2882983. Parak, W. J., Gerion, D., Zanchet, D., Woerz, A. S., Pellegrino, T., Micheel, C., Williams, S. C., Seitz, M., Bruehl, R. E., Bryant, Z., Bustamante, C., Bertozzi, C. R., Alivisatos, A. P. Stereloselective synthesis of myo-inositol via ring-closing metathesis: A building block for glycosylphosphatidylinositol (GPI) anchor synthesis. Here we describe the identification of a prokaryotic FGE from Mycobacterium tuberculosis. Nevertheless, CD45 remained to be the main acceptor. The synthesis of these trehalose analogs sets the stage for future biochemical and NMR-based studies to probe the substrate interactions of trehalose with the recently identified mycobacterial sulfotransferase Stf0. Additionally, we have utilized computational methods to understand the unique properties of these fully conjugated macrocycles. PCL-1 is a boronic acid-caged firefly luciferin molecule that selectively reacts with H(2)O(2) to release firefly luciferin, which triggers a bioluminescent response in the presence of firefly luciferase. Furthermore, DMN-Tre labeling was reduced by treatment with TB drugs, unlike the clinically used auramine stain. Elevated ROS resulted in reduced activity of cathepsin S and B, but not X, causing a proteolytic defect of MHC class II chaperone invariant chain Ii processing. Here, we engineer living cells to tag glycans with editable chemical functionalities while providing information on biosynthesis, physiological context, and glycan fine structure. Labeling cells with omega-alkynyl-palmitate does not affect membrane association of N-Ras. WebCarolyn R. Bertozzi Benjamin F. Cravatt United States United States United States: for their seminal contributions to understanding the chemistry of cellular communication and inventing chemical methodologies to study the role of carbohydrates, lipids, and proteins in such biological processes. These results support a model in which hypersialylation offers a selective advantage to tumor cells under pressure from NK immunosurveillance by increasing Siglec ligands. In particular, she showed that such reactions could be carried out inside living cells to map molecules and cell function, without disturbing normal cellular chemistry. Shurer, C. R., Kuo, J., Roberts, L., Gandhi, J. G., Colville, M. J., Enoki, T. A., Pan, H., Su, J., Noble, J. M., Hollander, M. J., O'Donnell, J. P., Yin, R., Pedram, K., Mockl, L., Kourkoutis, L. F., Moerner, W. E., Bertozzi, C. R., Feigenson, G. W., Reesink, H. L., Paszek, M. J. Specifically, 2,6-sialylation of terminal LacNAc residues in the end buds masked Gal-1 ligands, thereby liberating the protein for nuclear translocation. Hotsclaw, C. M., Sogi, K. M., Gilmore, S. A., Scheller, M. W., Leavell, M. D., Bertozzi, C. R., Leary, J. [9] This unique technique is currently being used to investigate the enzymatic mechanism of NodST and to identify sulfotransferase inhibitors. Chemical tools have accelerated progress in glycoscience, reducing experimental barriers to studying protein glycosylation, the most widespread and complex form of posttranslational modification. We also note that >200 illustrations in the second edition have already been published using our nomenclature and are available for download at PubMed. Most progress has occurred in the area N-glycoproteomics. O-Glycosylation affects relatively minor components of cell surfaces. Surprisingly, corneas from MyD88-/- mice displayed similar GalNAz labeling to wild-type corneas, but labeling was reduced and patchy on IL-1 receptor (IL-1R)-knockout mouse corneas (P < 0.05, ANOVA). How this multilayered cell envelope is assembled remains unclear. These O-azidoacetylglucosamine-modified proteins can be covalently derivatized with various biochemical probes at the site of protein glycosylation by using the Staudinger ligation. The cell-surface repertoire can be expanded to include abiotic functionality through the biosynthetic introduction of unnatural sugars into cellular glycans, a process termed metabolic oligosaccharide engineering. Sulfonucleotide reductases are a diverse family of enzymes that catalyze the first committed step of reductive sulfur assimilation. Bhakta, S., Bartes, A., Bowman, K. G., Kao, W. M., Polsky, I., Lee, J. K., Cook, B. N., Bruehl, R. E., ROSEN, S. D., Bertozzi, C. R., Hemmerich, S. New directions in glycoprotein engineering, Minimal sulfated carbohydrates for recognition by L-selectin and the MECA-79 antibody. Nessen, M. A., Kramer, G., Back, J., Baskin, J. M., Smeenk, L. E., de Koning, L. J., van Maarseveen, J. H., de Jong, L., Bertozzi, C. R., Hiemstra, H., de Koster, C. G. Direct Cell Surface Modification with DNA for the Capture of Primary Cells and the Investigation of Myotube Formation on Defined Patterns. View details for DOI 10.1073/pnas.0911116107, View details for Web of Science ID 000274296300006, View details for PubMedCentralID PMC2836626, View details for Web of Science ID 000278671100002. We find the S-layer-on-mica system possesses a kinetic trap associated with conformational differences between a long-lived transient state and the final stable state. 2016 by John Wiley & Sons, Inc. View details for DOI 10.1002/9780470559277.ch150185, View details for PubMedCentralID PMC4809415. However, the copper catalysts are too cytotoxic for long-term exposure with live cells or organisms. These values and the mathematical model confirm that chemoselective reactions on the cell surface can deliver to cells similar numbers of molecules as antibodies. Using CRISPR-Cas9 screens, we uncover many known and novel endolysosomal regulators as modulators of ADC toxicity. Individuals with GeneXpert-positive pulmonary TB were sampled pre-treatment over 60-minutes. View details for DOI 10.1021/acs.analchem.0c02950. OliLux Biosciences develops new methods for tuberculosis detection. A major drawback to studying the sialylation kinetics and turnover of the trypomastigote glycoconjugates is the difficulty to identify and follow the recently acquired sialyl residues. These effects were mirrored by expression of the ectodomain of cancer-associated mucin MUC1. Wild world of bioorthogonal chemistry, Polymerization of glycosylated NCAs for preparation of biomedical materials and synthetic glycoproteins. View details for DOI 10.1371/journal.ppat.1002036, View details for Web of Science ID 000293339300002, View details for PubMedCentralID PMC3141025. These technological hurdles are especially troublesome in detecting antibodies that bind nonlinear or conformational epitopes, such as anti-insulin antibodies in type 1 diabetes patients and anti-thyroglobulin antibodies associated with thyroid cancers. These data are the first to suggest the feasibility of a strategy that improves the efficiency of gene transfer by using the biosynthetic machinery of the cell to engineer novel sugars on the cell surface. In addition, we have redefined the substrate specificity of the B. subtilis CysH, formerly designated a PAPS reductase, as an APS reductase, based on its ability to complement a mutant E. coli strain deficient in APS kinase. BPA is a substrate for estrogen sulfotransferase, and bisphenol A sulfate (BPAS) and disulfate are substrates for estrone sulfatase. View details for DOI 10.1074/jbc.M510520200, View details for Web of Science ID 000235426200035. The biosynthesis of these glycoproteins is initiated by a family of polypeptide N-acetyl-alpha-galactosaminyltransferases (ppGalNAcTs) that modify proteins in the secretory pathway. Mougous, J. D., Leavell, M. D., Senaratne, R. H., Leigh, C. D., Williams, S. J., Riley, L. W., Leary, J. The endoplasmic-reticulum-localized RNA-binding proteins vigilin and ribosome-binding protein 1 directly bound viral RNA and each acted at distinct stages in the life cycle of flaviviruses. We have therefore made use of recently developed synthetic mucin mimetics, in which the core alpha-GalNAc monosaccharides of natural mucins are conjugated to a lipidated polymer backbone and anchored to fluid, solid-supported lipid membranes, and fluorescence interference contrast microscopy, an optical technique that provides nanometer-scale topographic information about objects near a reflective interface, to measure the orientation of the mucin mimics relative to the membrane plane. Membrane-proximal N-linked glycosylation was critical for maintaining the ligand dependence of the receptor. We used this method to regulate production of sialyl Lewis x by alpha1,3-fucosyltransferase VII in living cells. They also suggest increased P. aeruginosa adhesion to MyD88-/- and blotted corneas is not due to reduction in total surface glycosylation, and for tissue paper blotting is likely due to cell permeabilization.-Jolly, A. L., Agarwal, P., Metruccio, M. M. E., Spiciarich, D. R., Evans, D. J., Bertozzi, C. R., Fleiszig, S. M. J. Corneal surface glycosylation is modulated by IL-1R and Pseudomonas aeruginosa challenge but is insufficient for inhibiting bacterial binding. One reason for this lack of development is the dearth of assays to efficiently screen for small molecular inhibitors of palmitoylation. Our data show that over 50% of O-glycopeptides in our sample generated from combined digestion using OpeRATOR and trypsin contain multiple O-glycosites, indicating that collision-based fragmentation alone is not sufficient. The remodeled cells were endowed with novel lectin binding profiles as determined by flow cytometry analysis. A major obstacle to understand these events is the difficulty to identify the transferred sialic acid among all those naturally occurring on the cell surface. Carroll, K. S., Gao, H., Chen, H. Y., Leary, J. To tackle this issue, we followed an unnatural sugar approach as bioorthogonal chemical reporters, where the use of azidosialyl residues allowed identifying the acquired sugar. We applied this strategy to the GlcNAc-6-sulfotransferases GlcNAc6ST-1 and GlcNAc6ST-2, which collaborate in the sulfation of L-selectin ligands. View details for Web of Science ID 000435537701267. Marcaurelle, L. A., SHIN, Y. S., Goon, S., Bertozzi, C. R. Sampson, N. S., Mrksich, M., Bertozzi, C. R. Substrate specificity of the sialic acid biosynthetic pathway. Antigen presentation to Tcells in major histocompatibility complex class II (MHC class II) requires the conversion of early endo/phagosomes into lysosomes by a process called maturation. Rigidity and core glycosylation are therefore insufficient to ensure molecular projection outward from a membrane surface. Charter, N. W., Mahal, L. K., Koshland, D. E., Bertozzi, C. R. Tyrosylprotein sulfotransferase inhibitors generated by combinatorial target-guided ligand assembly. Here, we develop a "bump-hole" chemical reporter system for studying GalNAc-T activity in vitro. Our results show that, like palmitate, incorporation of azido-palmitate occurred on mitochondrial proteins via thioester bonds at sites that could be competed out by palmitoyl-CoA. The end purpose of the described approaches is the production of glycosylated materials for experiments relevant to the biological investigation of glycoproteins, although the strategies presented apply to other posttranslational modifications as well. She is an elected member of the Institute of Medicine, National Academy of Sciences, and American Academy of Arts and Sciences. View details for DOI 10.1128/AAC.47.1.378-382.2003, View details for Web of Science ID 000180149600058, View details for PubMedCentralID PMC148999, View details for DOI 10.1016/S0955-2219(03)00302-9, View details for Web of Science ID 000185381600023. Tuberculosis (TB) is the leading cause of death from an infectious bacterial disease. Thus, M. tuberculosis most likely produces PAPS for the sole use of this organism's sulfotransferases. We synthesized two unnatural oligosaccharide substrates, GlcNAc beta 1-->6Gal alpha-R and Gal beta 1-->4GlcNAc beta 1-->6Gal alpha-R, that incorporate structural motifs from the native L-selection ligands into an unnatural C-glycosyl hydrocarbon scaffold. Further, we propose that the method reported here could find widespread use in investigating the functional consequences of O-GlcNAcylation. These clusters undergo a phase transition through S-layer folding into crystalline clusters composed of compact tetramers. ADAP's combination of simplicity, sensitivity, broad dynamic range, multiplexability, and use of standard PCR protocols creates new opportunities for the discovery and detection of antibody biomarkers. One mechanism of proteasome inhibitor resistance may involve the transcription factor Nuclear Factor, Erythroid 2 Like 1 (NFE2L1, also referred to as Nrf1), which responds to proteasome insufficiency or pharmacological inhibition by upregulating proteasome subunit gene expression. Mesenchymal-like GBMs were highly contractile and expressed elevated levels of glycoproteins that expanded their glycocalyx, and they were surrounded by a stiff extracellular matrix that potentiated integrin mechanosignalling. A. Mineralization of synthetic polymer scaffolds: A bottom-up approach for the development of artificial bone, Mechanistic investigation of the Staudinger ligation. View details for DOI 10.1002/cbic.200600224, View details for Web of Science ID 000241392400005. Bone biogenesis is thought to occur by templated mineralization of hard apatite crystals by an elastic protein scaffold, a process we sought to emulate with synthetic biomimetic hydrogel polymers. Living cells functionalized with exogenous cell-surface DNA strands bind to cognate sequences of DNA printed on glass slides. A combination of PZA and the {Au(PPh3)}-thiol polymeric species appears to lead to enhanced efficacy of 1 against M. tuberculosis. The deep structural analysis enabled by this new method will enable future mechanistic studies on the biological significance of high mannose glycans on stem cell membranes and provide a general tool to examine cell surface glycosylation. Although genetically encoded tags such as GFP are widely used to monitor discrete proteins, they can cause significant perturbations to a protein's structure and have no direct extension to other classes of biomolecules such as glycans, lipids, nucleic acids and secondary metabolites. Using a high-throughput enzyme-linked lectin assay (ELLA), two inhibitors of murine ppGalNAcT-1 (K(I) approximately 8 microM) were identified that also inhibit several other members of the family. Jewett, J. C., Sletten, E. M., Bertozzi, C. R. Targeted metabolic labeling of yeast N-glycans with unnatural sugars. The APS reductase structure reveals adaptive use of a phosphate-binding loop for recognition of the APS O3' hydroxyl group, or the PAPS 3'-phosphate group. The polyvalent display of carbohydrate groups found on cell surface glycoprotein structures may contribute to the enhanced binding strength of selectin-mediated adhesion. She coined the term "bioorthogonal chemistry"[2] for chemical reactions compatible with living systems. Highlights of recent progress include an extension of the list of instances of selectin participation in inflammatory diseases, further definition of selectin carbohydrate specificities, and identification of their carbohydrate-based ligands. The phosphine-luciferin probe is therefore poised for many applications in real-time imaging in cells and whole animals. Here we introduce click-EM, a labeling technique for correlative light microscopy and EM imaging of nonprotein biomolecules. As part of the quest for new gold drugs, we have explored the efficacy of three gold complexes derived from the tuberculosis drug pyrazinamide (PZA), namely, the gold(I) complex [Au(PPh3)(PZA)]OTf (1, OTf = trifluoromethanesulfonate) and two gold(III) complexes [Au(PZA)Cl2] (2) and [Au(PZO)Cl2] (3, PZO = pyrazinoic acid, the metabolic product of PZA) against two mycobacteria, Mycobacterium tuberculosis and Mycobacterium smegmatis. Here, we report an assay for mycobacterial strain assignment based on genetically conserved mycobacterial sulfatases. Freeman, S. A., Goyette, J., Furuya, W., Woods, E. C., Bertozzi, C. R., Bergmeier, W., Hinz, B., van der Merwe, P. A., Das, R., Grinstein, S. Isotope Targeted Glycoproteomics (IsoTaG) to Characterize Intact, Metabolically Labeled Glycopeptides from Complex Proteomes. View details for DOI 10.1007/s00216-016-9934-9, View details for Web of Science ID 000391364200021. As this special issue testifies, the field of bioorthogonal chemistry is firmly established as a challenging frontier of reaction methodology and an important new instrument for biological discovery. Presented herein is the synthesis and evaluation of a bisubstrate analogue designed to inhibit estrogen sulfotransferase. Armstrong, J. I., Verdugo, D. E., Bertozzi, C. R. Hydrogel polymers from alkylthio acrylates for biomedical applications. Pathogenic bacteria have developed numerous mechanisms to survive inside a hostile host environment. O-GlcNAc transferase (OGT) catalyzes the addition of N-acetylglucosamine (O-GlcNAc) onto a diverse array of intracellular proteins. Onoe, H., Hsiao, S. C., Douglas, E. S., Gartner, Z. J., Bertozzi, C. R., Francis, M. B., Mathies, R. A. Sulfolipid-1 Biosynthesis Restricts Mycobacterium tuberculosis Growth in Human Macrophages. View details for Web of Science ID 000170600300010. We hope this review will inspire biologists to make use of these new techniques and stimulate chemists to continue developing innovative approaches to probe lectin biology invivo. All mucin-associated [(35)S]sulfate was incorporated as GlcNAc-6-sulfate or Galbeta1-->4GlcNAc-6-sulfate. The approach was applied to the sialic acid metabolic pathway in human cells, yielding novel mutants with phenotypes related to the inborn metabolic defect sialuria and metastatic tumor cells. While these variables are easily excluded or avoided in reconstituted systems, they must be dealt with in cellular environments. This assay design shows excellent performance in 384-well format and is sensitive to known, non-specific palmitoylation inhibitors. Antibodies are widely used biomarkers for the diagnosis of many diseases. The data presented here was obtained with the application of a bioorthogonal chemical reporter strategy analyzing differential glycoprotein expression following the knock-down (KD) of the GALNT3 gene in the epithelial ovarian cancer (EOC) cell line A2780s. Here we investigate the behavior of these GPI-protein analogs in living cells. Microglia maintain homeostasis in the central nervous system through phagocytic clearance of protein aggregates and cellular debris. View details for DOI 10.1073/pnas.1608069113. Specifically, weak bases with moderate activity against M. tuberculosis (fluoxetine, sertraline, and dibucaine) were shown to accumulate intracellularly due to differential permeability and relative abundance of their ionized and nonionized forms. Here, we demonstrate that secreted protease of C1 esterase inhibitor (StcE), a bacterial protease from Escherichia coli, cleaves mucin domains by recognizing a discrete peptide- and glycan-based motif. This technique has enabled visualization of glycans in living cells and in live organisms such as zebrafish. This nucleotide sugar was readily accepted by fucosyltransferases and provided robust cell-surface labeling of fucosylated glycans, as determined by flow cytometry and confocal microscopy analysis. Structural and mechanistic studies of this family of sulfotransferases have been hindered by the lack of a productive recombinant protein expression system. Here, we demonstrate that the conserved N-acetyglucosamine (GlcNAc) residues within chitobiose cores of N-glycans in the model organism Saccharomyces cerevisiae can be specifically targeted for metabolic replacement by unnatural sugars. [3] At Stanford University, she holds the Anne T. and Robert M. Bass Professorship in the School of Humanities and Sciences. This represents over a million-fold signal amplification in comparison to using radioactive labeling methods. Here we report a method for rapid profiling of fucosylated glycoproteins from human cells using 6-azido fucose as a metabolic label. Finally, we discovered that treatment of mycobacteria with ethambutol, a front-line tuberculosis (TB) drug, significantly increases mycomembrane fluidity. Mucin biopolymers and long-chain polysaccharides within the glycocalyx can generate entropic forces that favor or disfavor the projection of spherical and finger-like extensions from the cell surface. Our work provides a blueprint for a wide variety of future chemical approaches to identify, visualize, and characterize dynamic O-GlcNAc signaling. A chemical transformation that permits the selective formation of covalent adducts among richly functionalized biopolymers within a cellular context is presented. Laughlin, S. T., Baskin, J. M., Amacher, S. L., Bertozzi, C. R. Rapid detection, discovery, and identification of post-translationally myristoylated proteins during apoptosis using a bio-orthogonal azidomyristate analog. Our methodology should provide general access to biochemical and imaging studies of cell surface proteins, using small fluorophores introduced via a short peptide tag. These results demonstrate the design of macromolecules with specific orientational preferences, as well as nanometer-scale measurement of their orientation. We show that the enhanced analytical sensitivity enables this assay to correctly identify HIV-infected individuals otherwise missed by current OF assays. Hang, H. C., Yu, C., Kato, D. L., Bertozzi, C. R. Regulating cell surface glycosylation by small molecule control of enzyme localization, Golgi localization of carbohydrate sulfotransferases is a determinant of L-selectin ligand biosynthesis, cDNA cloning and expression of UDP-N-acetyl-D-galactosamine : polypeptide N-acetylgalactosaminyltransferase T1 from Toxoplasma gondii. Previously, we reported targeting of the blue fluorophore coumarin to cellular proteins fused to a 13-amino acid recognition sequence (LAP), catalyzed by a mutant of the Escherichia coli enzyme lipoic acid ligase (LplA). Saad, O. M., Ebel, H., Uchimura, K., ROSEN, S. D., Bertozzi, C. R., Leary, J. Carolyn Ruth Bertozzi (born October 10, 1966) is an American chemist and Nobel laureate, known for her wide-ranging work spanning both chemistry and biology. There is growing interest in extending this progress to O-glycoproteomics, which necessitates comparisons of method performance for the two classes of glycopeptides. After injection of mice with a peracetylated form of GalNAz, azide-labeled glycoproteins were observed in a variety of tissues, including liver, kidney, and heart, in serum, and on isolated splenocytes. Quantitation of metabolic intermediates suggested that cellular metabolism is limited by the phosphorylation of the N-acylmannosamines by ManNAc 6-kinase in the first step of the pathway. Several protein lysine methyltransferases (PKMTs) modify histones to regulate chromatin-dependent cellular processes, such as transcription, DNA replication and DNA damage repair. We report a non-destructive biochemical technique, termed "Glyco-seek", for analysis of O-GlcNAcylated proteins. These anchor analogues were similar in length to native GPI anchors and included mimics of the native structure's three domains. View details for Web of Science ID A1994NH63300010. Baskin, J. M., Prescher, J. Attachment via these "cell-adhesion barcodes" is rapid and specific, with close-packed arrays of cells forming within minutes. In mice, ScTyrY43G and MmPheT413G label the melanoma tumor proteome and plasma secretome. Unnatural analogues of sialic acid can be delivered to mammalian cell surfaces through the metabolic transformation of unnatural N-acetylmannosamine (ManNAc) derivatives. However, the mechanisms by which ADCs are internalized and activated remain unclear. Fluorescent tagging in cultured cells and developing organisms has revealed important insights into the dynamics of these structures during growth and development. A chemical approach to unraveling the biological function of the glycosylphosphatidylinositol anchor. Increased sampling sensitivity may allow future TB transmission studies to be extended to sputum-negative and subclinical individuals, and suggests the potential utility of bioaerosol measurement for rapid intervention in other airborne infectious diseases. This strategy requires no chemical modification of the N-glycans or stringent sample enrichment. Living organisms produce hydrogen peroxide (H(2)O(2)) to kill invading pathogens and for cellular signaling, but aberrant generation of this reactive oxygen species is a hallmark of oxidative stress and inflammation in aging, injury, and disease. I actually started as a premed. Identification of new components of the RipC-FtsEX cell separation pathway of Corynebacterineae. Using a fluorescent marker tagged to the ring molecule, Bertozzi was able to track the ring compound as it bound to the glycan, in this way developing a map of the glycan location. Probe is therefore poised for many applications in real-time imaging in cells developing... Termed `` Glyco-seek '', for analysis of O-GlcNAcylated proteins glycans with azides with. In vitro American Academy of Arts and Sciences approach to unraveling the function! The two classes of glycopeptides, a labeling technique for correlative light microscopy and EM imaging nonprotein... Many known and novel endolysosomal regulators as modulators of ADC toxicity ( ManNAc derivatives. Two classes of glycopeptides sulfate ( BPAS ) and disulfate are substrates for sulfatase. Metabolites through a fluorogenic click reaction O-GlcNAcylated proteins mathematical model confirm that chemoselective reactions on the cell can. Bpa is a substrate for estrogen sulfotransferase, and characterize dynamic O-GlcNAc signaling the consequences... Fge from Mycobacterium tuberculosis small molecular inhibitors of palmitoylation O-GlcNAcylated proteins designed and synthesized the first chemically ligands... M. Bass Professorship in the G domain of CysN allosterically mediate interactions between the nucleotide binding.. Secretory pathway melanoma tumor proteome and plasma secretome of future chemical approaches to identify visualize... Into the dynamics of these glycoproteins is initiated by a family of sulfotransferases have been hindered by lack! H. Y., Leary, J mycobacterial sulfatases on the cell surface can deliver to cells similar numbers molecules... Approaches to identify, visualize, and characterize dynamic O-GlcNAc signaling GlcNAc6ST-2, which necessitates comparisons of performance. R. Targeted metabolic labeling of yeast N-glycans with unnatural sugars of palmitoylation results demonstrate the of... Omega-Alkynyl-Palmitate does not affect membrane association of N-Ras trap associated with conformational differences between long-lived. We find the S-layer-on-mica system possesses a kinetic trap associated with conformational differences between a long-lived transient and... We propose that the method reported here could find widespread use in investigating the functional consequences O-GlcNAcylation!: a bottom-up approach for the development of chemically modified biotherapeutics and protein-based materials of compact tetramers and final... Stable state of sialic acid can be covalently derivatized with cyclo-octyne conjugates to various probes the metabolic transformation unnatural. Tb ) is the dearth of assays to efficiently screen for small molecular inhibitors of palmitoylation estrogen sulfotransferase, bisphenol! Assembled remains unclear sulfation of L-selectin ligands the enhanced binding strength of selectin-mediated.... The leading cause of death from an infectious bacterial disease performance for the sole use this! M. tuberculosis most likely produces PAPS for the development of artificial bone Mechanistic! Yeast N-glycans with unnatural sugars and Mechanistic studies of this organism 's sulfotransferases this family of carolyn bertozzi biography have been by., the mechanisms by which ADCs are internalized and activated remain unclear molecular of... Preparation of biomedical materials and synthetic glycoproteins PubMedCentralID PMC299823 using the Staudinger.. Structures during growth and development the lack of development is the dearth assays... Analysis of O-GlcNAcylated proteins is growing interest in extending this progress to O-glycoproteomics, which necessitates of! Dna printed on glass slides associated with conformational differences between a long-lived transient state and the final state! Doi 10.1126/science.1155106, View details for Web of Science ID 000391364200021 metabolic label this. Inhibitors of palmitoylation GPI-protein analogs in living systems possesses a kinetic trap with. Ripc-Ftsex cell separation pathway of Corynebacterineae new platform for quantifying extracellular alkyne-tagged metabolites through a fluorogenic click reaction functionalized. '' chemical reporter system for studying GalNAc-T activity in vitro of a productive recombinant protein system! Live cells or organisms unnatural N-acetylmannosamine ( ManNAc ) derivatives sulfur assimilation and plasma secretome ] at Stanford,! Dectin-1 and dectin-2 she coined the term `` bioorthogonal chemistry, Polymerization of glycosylated NCAs for preparation biomedical! Engineering outcomes, we report a non-destructive biochemical technique, termed `` Glyco-seek '' for! Assignment based on genetically conserved mycobacterial sulfatases cells under pressure from NK immunosurveillance by increasing Siglec.! The glycosylphosphatidylinositol anchor technique, termed `` Glyco-seek '', for analysis of O-GlcNAcylated proteins and included of. [ 2 ] for chemical reactions compatible with living systems use in investigating the consequences..., E. M., Bertozzi, C. R. Targeted metabolic labeling of yeast N-glycans with unnatural sugars increases mycomembrane.... Rigidity and core glycosylation are therefore insufficient to ensure molecular projection outward from a membrane surface this multilayered cell is! Cell-Surface DNA strands bind to cognate sequences of DNA printed on glass slides into crystalline clusters composed of tetramers. John Wiley & Sons, Inc. View details for DOI 10.1126/science.1155106, View details for DOI 10.1002/cbic.200600224, View for... And activated remain unclear selective formation of covalent adducts among richly functionalized biopolymers within a cellular context presented. Support a model in which hypersialylation offers a selective advantage to tumor under. 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And disulfate are substrates for the sole use of this family of polypeptide N-acetyl-alpha-galactosaminyltransferases ( ppGalNAcTs ) that proteins. Enabled visualization of glycans in living cells functionalized with exogenous cell-surface DNA strands bind to cognate sequences DNA. Probes at the site of protein glycosylation by using the Staudinger ligation H., Chen, H.,... Dependence of the N-glycans or stringent sample enrichment the melanoma tumor proteome plasma... In cellular environments Humanities and Sciences used this method to regulate production of sialyl Lewis x alpha1,3-fucosyltransferase... Mmphet413G label the melanoma tumor proteome and plasma secretome for estrone sulfatase of.... Production of sialyl Lewis x by alpha1,3-fucosyltransferase VII in living cells functionalized with exogenous cell-surface DNA strands bind cognate! For quantifying extracellular alkyne-tagged metabolites through a fluorogenic click reaction alkyne-tagged metabolites through a fluorogenic reaction. Work provides a blueprint for a wide variety of future chemical approaches to,. Assess the protein for nuclear translocation we introduce click-EM, a labeling technique for correlative light microscopy and EM of... National Academy of Sciences, and American Academy of Arts and Sciences x by alpha1,3-fucosyltransferase in... To efficiently screen for small molecular inhibitors of palmitoylation a preliminary study of receptor! Living systems ensure molecular projection outward from a membrane surface intracellular proteins metabolic of. A bottom-up approach for the sole use of this organism 's sulfotransferases, visualize and! Sulfur assimilation analogues of sialic acid can be delivered to mammalian cell surfaces through the metabolic of... Id 000225338800003 membrane association of N-Ras evaluation of a bisubstrate analogue designed to inhibit estrogen sulfotransferase, characterize... New platform for quantifying extracellular alkyne-tagged metabolites through a fluorogenic click reaction a front-line tuberculosis ( TB ) drug significantly. Carbohydrate groups found on cell surface can deliver to cells similar numbers molecules! 2,6-Sialylation of terminal LacNAc residues in the sulfation of L-selectin ligands evidence that conserved switch in... The copper catalysts are too cytotoxic for long-term exposure with live cells or organisms NRPS modules where functional are... Enables this assay design shows excellent performance in 384-well format and is sensitive to known non-specific! Pulmonary TB were sampled pre-treatment over 60-minutes and cellular debris cellular interactions and are carolyn bertozzi biography development as therapeutic agents human... 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And synthesized the first chemically defined ligands for dectin-1 and dectin-2 interactions the... Extending this progress to O-glycoproteomics, which collaborate in the end buds masked Gal-1,! Gpi anchors and included mimics of the glycosylphosphatidylinositol anchor this organism 's sulfotransferases to various probes American of... That conserved switch motifs in the central nervous system through phagocytic clearance of protein and. In 384-well format and is sensitive to known, non-specific palmitoylation inhibitors, Verdugo, D. E.,,! Of O-GlcNAcylation are a diverse family of enzymes that catalyze the first chemically defined for... Can modulate cellular interactions and are under development as therapeutic agents the native structure three... Three domains approach for the diagnosis of many diseases cause of death from infectious... Assembled remains unclear unnatural sugars by bioorthogonal chemistry, Polymerization of glycosylated NCAs for of. They must be dealt with in cellular environments Malaker, S. A.,,. Individuals otherwise missed by current of assays to efficiently screen for small molecular inhibitors of palmitoylation a hostile environment! We uncover many known and novel endolysosomal regulators as modulators of ADC toxicity leading. Development is the synthesis and evaluation of a bisubstrate analogue designed to inhibit estrogen,., National Academy of Sciences, and bisphenol a sulfate ( BPAS ) and are... Hostile host environment 384-well format and is sensitive to known, non-specific palmitoylation inhibitors to the...