Metabolic Glycan Engineering in Live Animals: Using Bio‐orthogonal Chemistry to Alter Cell Surface Glycans

Dube, Danielle H.; Williams, DA

doi:10.1002/9783527344406.ch8

Cited by 6 publications

(4 citation statements)

References 115 publications

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“…Azide-containing sugars have facilitated tagging of labeled glycans with bioorthogonal reaction partners (e.g., alkynes, phosphines) to enable detection, identification, and visualization of bacterial glycans in live cells and animal infection models. 6,40,42 Motivated by the observation that azide-containing sugars provide a readout of bacterial glycan biosynthesis, we reasoned that metabolic substrates could effectively report on glycan biosynthesis defects and guide the identification of genes required for glycan biosynthesis, even in the absence of detailed pathway or glycan information. Guided by previous work in which azide-containing sugars led to the visualization of the full complement of glycoproteins synthesized by bacteria (Figure 1B), 37,38 we hypothesized that the disruption of genes involved in glycoprotein biosynthesis would yield a defective glycoprotein biosynthesis "fingerprint" relative to wild-type bacteria (Figure 1C).…”

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“…Since the initial demonstration that bacterial cells supplemented with unnatural monosaccharides bearing azides take up those sugars, process them via endogenous carbohydrate biosynthetic pathways, and install them into cellular glycans in place of natural monosaccharides (Figure B), MOE has led to the discovery, − tracking, and inhibition of bacterial glycosylation systems. Azide-containing sugars have facilitated tagging of labeled glycans with bioorthogonal reaction partners (e.g., alkynes, phosphines) to enable detection, identification, and visualization of bacterial glycans in live cells and animal infection models. ,, …”

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Metabolic Glycan Labeling-Based Screen to Identify Bacterial Glycosylation Genes

et al. 2020

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Bacterial cell surface glycans are quintessential drug targets due to their critical role in colonization of the host, pathogen survival, and immune evasion. The dense cell envelope glycocalyx contains distinctive monosaccharides that are stitched together into higher order glycans to yield exclusively bacterial structures that are critical for strain fitness and pathogenesis. However, the systematic study and inhibition of bacterial glycosylation enzymes remains challenging. Bacteria produce glycans containing rare sugars refractory to traditional glycan analysis, complicating the study of bacterial glycans and the identification of their biosynthesis machinery. To ease the study of bacterial glycans in the absence of detailed structural information, we used metabolic glycan labeling to detect changes in glycan biosynthesis. Here, we screened wild-type versus mutant strains of the gastric pathogen Helicobacter pylori, ultimately permitting the identification of genes involved in glycoprotein and lipopolysaccharide biosynthesis. Our findings provide the first evidence that H. pylori protein glycosylation proceeds via a lipid carrier-mediated pathway that overlaps with lipopolysaccharide biosynthesis. Protein glycosylation mutants displayed fitness defects consistent with those induced by small molecule glycosylation inhibitors. Broadly, our results suggest a facile approach to screen for bacterial glycosylation genes and gain insight into their biosynthesis and functional importance, even in the absence of glycan structural information.

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Metabolic Glycan Labeling-Based Screen to Identify Bacterial Glycosylation Genes

et al. 2020

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“…Azide-labeled carbohydrates are endocytosed by cells and integrated with glycan biosynthesis in various glycoconjugates. The cells are incubated for periods of 24 to 72 hours to allow the synthesis of surface glycoproteins to be monitored [11,12].…”

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Can PDT Alter the Glycosylation of the Tumor Cell Membrane?

Godoi¹,

Ferreira‐Strixino²,

Silva³

et al. 2021

Photodynamic Therapy - From Basic Science to Clinical Research

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Photodynamic Therapy (PDT) is a cancer treatment that used the interaction of a photosensitizing drug and a light source. PDT can lead to changes in the expression of various cellular elements, compromising cell adhesion, and cytoskeleton integrity in cells undergoing treatment. However, the pathways of cellular alterations caused by this treatment are little known. Alterations in expression in surface glycoproteins and glycolipids are significant features in malignant tumor transformation and are strongly associated with tumor cell adhesion, invasion, and metastasis. This study evaluated photodynamic therapy effects on indirect distribution surface glycoproteins in human laryngeal carcinoma HEp-2 cell line surface, using Click-iT™ Metabolic Glycoprotein Labeling Reagent. Aluminum Phthalocyanine Tetrasulfonate (AlPcS4) was administrated at 5 μM/mL, followed by one hour of the incubation period for its accumulation in the tumor cells. After this time, cultures were irradiated with LED (light-emitting diode) dispositive (BioPdi/IRRAD-LED) λ = 660 nm. Evaluation of glycoproteins was performed by flow cytometry. Knowledge of the cellular alterations caused by the treatment will allow obtaining tools for the potentiation or optimization and personalization of the anticancer treatment. This therapy has a low cost and better efficacy, when applied early, about radiotherapy chemotherapy.

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“…Metabolic oligosaccharide engineering (MOE) has emerged as a powerful method for the efficient investigation of bacterial glycans. , Originally pioneered by Bertozzi, , Reutter and colleagues , for studying glycans in mammalian systems, MOE is a two-step chemical approach that has been adapted to probe the bacterial glycome (Figure A). , Briefly, bacteria treated with azide-containing sugar analogues remodel their glycocalyx to yield metabolically labeled cellular glycans. Then, bioorthogonal chemistry , is used to produce detectable signals that aid the tracking, enrichment, and characterization of bacterial glycans.…”

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Synthesis and Application of Rare Deoxy Amino l-Sugar Analogues to Probe Glycans in Pathogenic Bacteria

et al. 2022

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Bacterial cell envelope glycans are compelling antibiotic targets as they are critical for strain fitness and pathogenesis yet are virtually absent from human cells. However, systematic study and perturbation of bacterial glycans remains challenging due to their utilization of rare deoxy amino L-sugars, which impede traditional glycan analysis and are not readily available from natural sources. The development of chemical tools to study bacterial glycans is a crucial step toward understanding and altering these biomolecules. Here we report an expedient methodology to access azide-containing analogues of a variety of unusual deoxy amino L-sugars starting from readily available L-rhamnose and L-fucose. Azidecontaining L-sugar analogues facilitated metabolic profiling of bacterial glycans in a range of Gram-negative bacteria and revealed differential utilization of L-sugars in symbiotic versus pathogenic bacteria. Further application of these probes will refine our knowledge of the glycan repertoire in diverse bacteria and aid in the design of novel antibiotics.

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Metabolic Glycan Engineering in Live Animals: Using Bio‐orthogonal Chemistry to Alter Cell Surface Glycans

Cited by 6 publications

References 115 publications

Metabolic Glycan Labeling-Based Screen to Identify Bacterial Glycosylation Genes

Metabolic Glycan Labeling-Based Screen to Identify Bacterial Glycosylation Genes

Can PDT Alter the Glycosylation of the Tumor Cell Membrane?

Synthesis and Application of Rare Deoxy Amino l-Sugar Analogues to Probe Glycans in Pathogenic Bacteria

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