O-Linked -N-acetylgalactosamine (O-GalNAc) glycans constitute a major part of the human glycome. Northern blot analysis identified a single 5.5-kb ppGalNAc-T transcript. View details for DOI 10.1016/j.devcel.2019.04.035. Polysialyltransferases catalyze the glycosylation of the neural cell adhesion molecule (NCAM) with polysialic acid (PSA). Azides installed within cell surface glycoconjugates by metabolism of a synthetic azidosugar were reacted with a biotinylated triarylphosphine to produce stable cell-surface adducts. Palaniappan, K. K., Pitcher, A. View details for DOI 10.1073/pnas.1201504109, View details for Web of Science ID 000307551700033, View details for PubMedCentralID PMC3420203. Moeckl, L., Pedram, K., Roy, A., Bertozzi, C., Moerner, W. Aebersold, R. n., Agar, J. N., Amster, I. J., Baker, M. S., Bertozzi, C. R., Boja, E. S., Costello, C. E., Cravatt, B. F., Fenselau, C. n., Garcia, B. Potential for therapeutics and diagnostics, Kinetic measurements and mechanism determination of Stf0 sulfotransferase using mass spectrometry. A Fourier transform ion cyclotron resonance mass spectrometer provided accurate mass measurements, and a quadrupole time-of-flight instrument was used to obtain characteristic fragment ions and partial carbohydrate sequences. The primary sequence of APS reductase is distinguished by a conserved iron-sulfur cluster motif, -CC-X( approximately )(80)-CXXC-. They originate from biosynthetic pathways comprising an assembly line of glycosyltransferases within the Golgi compartment. Sulfite is then released in a thioredoxin-dependent manner. We demonstrated that the kinetic parameters of the assembly process depend on DNA sequence complexity, density, and total cell concentration. The prokaryotic homolog exhibits remarkable structural similarity to human FGE, including the position of catalytic cysteine residues. Random mutagenesis yielded the best JamB variant with 20-fold increased activity in E. coli. and Irmgard Chu Distinguished Professorship in Chemistry (2005); Havinga Medal, Univ. These reactions possess extreme selectivity and biocompatibility, such that their participating reagents can form covalent bonds within richly functionalized biological systems--in some cases, living organisms. A strain-promoted [3+2] azide-alkyne cycloaddition for covalent modification of blomolecules in living systems. She has found a smart way to connect cells together. Previously, we described the biochemical activity of the sulfotransferase Stf0 that initiates SL-1 biosynthesis. Approximately one thousand proteins are annotated as being palmitoylated, and for some of these, including several oncogenes of the Ras and Src families, palmitoylation is indispensable for protein function. All mucin-associated [(35)S]sulfate was incorporated as GlcNAc-6-sulfate or Galbeta1-->4GlcNAc-6-sulfate. Glioblastoma multiforme (GBMs) are recurrent lethal brain tumours. 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. The controlled addition of structurally defined components to live cell membranes can facilitate the molecular level analysis of cell surface phenomena. The kinetics of this reaction are of paramount importance for studies of dynamic processes, particularly in living subjects. View details for Web of Science ID 000280227700012, View details for PubMedCentralID PMC2907714. This technique has enabled visualization of glycans in living cells and in live organisms such as zebrafish. We used these tools to perform the first site-directed mutagenesis study of a member of this sulfotransferase family, GST2. WebCarolyn Ruth Bertozzi (born October 10, 1966) is an American chemist. Using azide-derivatized analogues of fucose, we metabolically labeled cell-surface glycans and then detected the incorporated azides via copper-free click chemistry with a difluorinated cyclooctyne probe. Wojczyk, B. S., Stwora-Wojczyk, M. M., Hagen, F. K., Striepen, B., Hang, H. C., Bertozzi, C. R., Roos, D. S., Spitalnik, S. L. Metabolic oligosaccharide engineering as a tool for glycobiology, Formation of 1,1-alpha,alpha-glycosidic bonds by intramolecular aglycone delivery. Trypanosoma cruzi, the flagellate protozoan agent of Chagas disease or American trypanosomiasis, is unable to synthesize sialic acids de novo. View details for Web of Science ID 000237590400012. Most clinically approved biomarkers of cancer are glycoproteins, and those residing on the cell surface are of particular interest in biotherapeutics. Their dense glycosylation is believed to confer structural rigidity as well as molecular extension beyond the glycocalyx, crucial to interaction with the cellular environment. The reaction features a large dynamic range of reactivity, showcasing second-order rate constants as high as 2.310(3) M(-1) s(-1) using diboron reaction partners. Click chemistry and bio-orthogonal chemistry. The isolation of this antibody signals the potential of phage antibody libraries in the derivation of reagents specific for post-translational modifications, although the extensive screening required indicates that such antibodies are extremely rare. We characterized several proteins in LBC phagosomes that change in abundance on induction of autophagy, a process that has been previously implicated in the host defense against microbial pathogens. View details for PubMedCentralID PMC5985656. The quadricyclane ligation joins a small but growing list of tools for the selective covalent modification of biomolecules. This system thus constitutes an AND-type molecular logic gate that reports on the simultaneous presence of H(2)O(2) and caspase 8 activity. Through this approach, we achieved glycopolypeptides with high molecular weights and low dispersities. Herein we report a semisynthetic method of producing membrane-anchored proteins. Baskin, J. M., Dehnert, K. W., Laughlin, S. T., Amacher, S. L., Bertozzi, C. R. Rapid and selective detection of fatty acylated proteins using omega-alkynyl-fatty acids and click chemistry. The objective of these methods is to make glycoconjugate synthesis accessible to a broader community, thereby accelerating progress in glycobiology. These findings functionally link mucinous proteins with tumor aggression, and offer a new view of the cancer glycocalyx as a major driver of disease progression. An important class of carbohydrates studied within the field of glycobiology, heparin and heparan sulfate (HS) have been implicated in a diverse array of biological functions. However, changes in the glycan structure significantly affected membrane mobility, with the loss of monosaccharide units correlating to decreased diffusion. Synthetic Riboswitches That Induce Gene Expression in Diverse Bacterial Species. Secretory products can both diffuse, although very slowly, through the meshwork pores and interact noncovalently with the matrix. In previous studies, mannosamine analogues bearing simple N-acyl groups up to five carbon atoms in length were recognized as substrates by the biosynthetic machinery and transformed into cell surface sialoglycoconjugates [Keppler, O. T., et al. Their diffusion properties mirrored those of many natural membrane-associated biomolecules. Remodeling the sialylation status of cancer cells affected the susceptibility to NK cell cytotoxicity via Siglec-7 engagement in a variety of tumor types. Here we explore an alternative bioorthogonal reaction, the 1,3-dipolar cycloaddition of azides and cyclooctynes, also known as "Cu-free click chemistry," for labeling biomolecules in live mice. IsoStamp has the potential to enhance any proteomics platform that employs chemical labeling for targeted protein identification, including isotope coded affinity tagging, isobaric tagging for relative and absolute quantitation, and chemical tagging strategies for posttranslational modification. [47] Lycia Therapeutics focuses on developing technology which utilizes lysosome-targeting chimeras (LYTACs). Swarts, B. M., Holsclaw, C. M., Jewett, J. C., Alber, M., Fox, D. M., Siegrist, M. S., Leary, J. Liposomes displaying 3'-sulfo Lewis(X)-like oligosaccharides, on the other hand, show slight loss of binding with introduction of additional anionic functional groups for E- and P-selectin and negligible change for L-selectin. Our results reveal that these transferases have unique peptide and glycopeptide preferences demonstrating their substrate diversity and their likely roles ranging from initiating transferases to filling-in transferases. Importantly, glycopolymers containing biologically relevant branched oligosaccharides, such as sialyl Lewis(x), as well as sulfated glycosaminoglycan-like epitopes can be readily prepared using our methodology. View details for DOI 10.1074/jbc.M212127200, View details for Web of Science ID 000181466800038, View details for Web of Science ID 000181517000014. The utility of this approach is demonstrated through the observation of patterned cells as they communicate by diffusion-based paracrine signaling. Previous structural analysis of the oligosaccharide chains on GlyCAM-1, an endothelial-derived ligand for L-selectin, revealed two novel structures: 6'-sulfo sialyl Lewis x and 6-sulfo sialyl Lewis x. Here we report the design, synthesis, and in vivo applications of Peroxy Caged Luciferin-1 (PCL-1), a chemoselective bioluminescent probe for the real-time detection of H(2)O(2) within living animals. An essential step in this pathway is the activation of sulfate through adenylation by the enzyme ATP sulfurylase (ATPS), forming adenosine 5'-phosphosulfate (APS). We developed a novel universal MHC class II presentation assay based on a bio-orthogonal "clickable" antigen and showed that MHC class II presentation was disrupted by the inhibition of PIKfyve, which in turn resulted in reduced activation of CD4+ Tcells. These included proteins involved in the immune response, oxidation and reduction, and vesicle transport, as well as other cellular processes. EM has long been the main technique for imaging cell structures with nanometer resolution but has lagged behind light microscopy in the crucial ability to make specific molecules stand out. Furthermore, GalNAz-labeled B cells could be selectively targeted with a phosphine probe by Staudinger ligation within the living animal. These collective results provide a detailed mechanistic framework for understanding why nature chose this structurally unique monocopper active site to catalyze oxidase chemistry for sulfatase activation. Photoacoustic calorimetry combined with established absorption and fluorescence methodologies provides a complete arsenal for characterizing the photophysical properties of many systems. Genomic and proteomic profiling form a basis for biological discovery. Notably, PapA3 was selective for trehalose; no activity was observed with other structurally related disaccharides. Assimilatory 3'-phosphoadenosine 5'-phosphosulfate (PAPS) reductases are evolutionarily related, homologous enzymes that catalyze the same overall reaction, but do so in the absence of an [Fe-S] cluster. Fucosylated glycoproteins are involved in many cell-cell recognition events and are markers of embryonic and malignant tissue. An, H. J., Gip, P., Kim, J., Wu, S., Park, K. W., McVaugh, C. T., Schaffer, D. V., Bertozzi, C. R., Lebrilla, C. B. Metabolic labeling enables selective photocrosslinking of O-GlcNAc-modified proteins to their binding partners. Bertozzi has applied the techniques of bioorthogonal chemistry to study glycocalyx, the sugars that surround the cell membrane. Insects protect themselves against bacterial infection by secreting a battery of antimicrobial peptides into the hemolymph. We observed crosslinking between FG-repeat nucleoporins and nuclear transport factors, suggesting that O-GlcNAc residues are intimately associated with essential recognition events in nuclear transport. In conclusion, the new ADAP assay can reliably detect the three cardinal islet autoantibodies/antibodies in 1muL serum with high sensitivity. In the latter case, we trace increased height to an effector gene, GALNT7. Together, these studies show that two conserved sequence motifs, CCXXRKXXPL and SXGCXXCT, found in the C termini of all APS reductases, but not in PAPS reductases, may be used to predict the substrate specificity of these enzymes. However, little is known about how alterations in O-GlcNAc cycling affect human embryonic stem cell (hESC) neural differentiation. Our approach capitalizes on two features shared by these enzymes: their requirement of Golgi localization for activity on cellular substrates and the modularity of their catalytic and localization domains. Here, we describe how pH-dependent partitioning drives asymmetric antimicrobial drug distribution in M. tuberculosis-infected macrophages. Furthermore, the membrane-bound glycopolymers were internalized into early endosomes similarly to endogenous membrane components and were capable of specific interactions with protein receptors. Baker Family Director of Stanford ChEM-H, Anne T. and Robert M. Bass Professor in the School of Humanities and Sciences and Professor, by courtesy, of Chemical and Systems Biology and of Radiology, AB, Harvard University, Chemistry (1988). The artificial receptor enhanced adenoviral vector binding and gene transfer to cells that are relatively resistant to adenovirus infection. Dr. Carolyn R. Bertozzi Ph.D. serves as Independent Director of the Company. The cysH mutant was also susceptible to peroxynitrite and hydrogen peroxide in vitro. The original goal was to discuss the value of creating centers of excellence for training the next generation of biomedical investigators in the glycosciences. 1-68A and a 2-dehydroxy analogue, 1-68Aa, inhibit several purified LpxC orthologues. View details for Web of Science ID 000338992200029. 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. A., Kalscheuer, R., Bertozzi, C. R. Density Variant Glycan Microarray for Evaluating Cross-Linking of Mucin-like Glycoconjugates by Lectins, Direct observation of kinetic traps associated with structural transformations leading to multiple pathways of S-layer assembly. Unbound particles randomize direction by Brownian rotation too quickly to be detected. Trypanosoma cruzi, the agent of the American trypanosomiasis or Chagas disease, bypasses its lack of de novo synthesis of sialic acids by expressing a surface-anchored trans-sialidase. Here we report that H. ducreyi can use unnatural sialic acids bearing elongated N-acyl groups from three to seven carbon atoms in length, resulting in outer membrane presentation of unnatural sialyl-LOS. Here we report a new biocompatible CID, Tmp-SLF, which dimerizes E. coli DHFR and FKBP and has no endogenous mammalian targets that would lead to unwanted in vivo side effects. Mass spectrometry assays allowed us to identify other acceptors, mainly integrins. Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis, is a highly evolved human pathogen characterized by its formidable cell wall. The use of bioorthogonal chemistry has contributed to advances in different areas of biomedical research, including cancer drug development and molecular imaging. 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