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| Open AccessA universal glycoenzyme biosynthesis pipeline that enables efficient cell-free remodeling of glycans
Access to glycoenzymes for basic and applied research is limited by difficulties with their recombinant expression. Here, the authors describe a universal strategy for converting membrane-bound glycosyltransferases into water-soluble biocatalysts, which are expressed at high levels with retention of activity.
- Thapakorn Jaroentomeechai
- , Yong Hyun Kwon
- & Matthew P. DeLisa
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Article
| Open AccessThe retaining β-Kdo glycosyltransferase WbbB uses a double-displacement mechanism with an intermediate adduct rearrangement step
WbbB is a structurally unusual retaining glycosyltransferase. Here, the authors show that WbbB forms an Asp232-Kdo adduct prior to transfer to the saccharide acceptor. Therefore, unlike any previously studied glycosyltransferase, WbbB uses the double-displacement mechanism first proposed in 1953.
- Taylor J. B. Forrester
- , Olga G. Ovchinnikova
- & Matthew S. Kimber
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Article
| Open AccessMolecular basis for polysaccharide recognition and modulated ATP hydrolysis by the O antigen ABC transporter
Gram negative microbes strategically equip lipopolysaccharides with O antigens to expand the extracellular envelope. Here, the authors examine how the O antigen is captured by a special class of ABC transporters containing a carbohydrate-binding module accessory.
- Nicholas Spellmon
- , Artur Muszyński
- & Jochen Zimmer
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Article
| Open AccessReformation of the chondroitin sulfate glycocalyx enables progression of AR-independent prostate cancer
Chondroitin sulfate (CS) is one of the most abundant glycosaminoglycans in prostate cancers. Here the authors show that inhibition of the androgen receptor pathway leads to the upregulation of CS, which promotes prostate cancer growth and metastasis.
- Nader Al-Nakouzi
- , Chris Kedong Wang
- & Mads Daugaard
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Article
| Open AccessTryptophan C-mannosylation is critical for Plasmodium falciparum transmission
Here, Lopaticki et al. show that Plasmodium falciparum expresses a Dpy19 C-mannosyltransferase in the endoplasmic reticulum that glycosylates TSR domains. Functional characterization shows that PfDpy19 plays a critical role in transmission through mosquitoes as PfDpy19-deficiency abolishes C-glycosylation and destabilizes proteins relevant for gametogenesis and oocyst formation.
- Sash Lopaticki
- , Robyn McConville
- & Justin A. Boddey
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Article
| Open AccessCell surface glycan engineering reveals that matriglycan alone can recapitulate dystroglycan binding and function
Matriglycan, a repeating disaccharide on α-dystroglycan, is the receptor for Lassa virus and specific extracellular matrix proteins. Here, the authors demonstrate that matriglycan, in a length-dependent tunable manner, is both necessary and sufficient for protein binding and viral infection.
- M. Osman Sheikh
- , Chantelle J. Capicciotti
- & Geert-Jan Boons
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Article
| Open AccessEngineering substrate specificity of HAD phosphatases and multienzyme systems development for the thermodynamic-driven manufacturing sugars
Haloacid dehalogenase-like phosphatases are widespread across all domains of life and play a crucial role in the regulation of levels of sugar phosphate metabolites in cells. The authors report on the structure-guided engineering of phosphatases for dedicated substrate specificity for the conversion of sucrose and starch into fructose and mannose.
- Chaoyu Tian
- , Jiangang Yang
- & Yanhe Ma
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Article
| Open AccessRevealing the human mucinome
Mucin-domain glycoproteins are densely O-glycosylated proteins with unique secondary structure that imparts a large influence on cellular environments. Here, the authors develop a technique to selectively enrich and characterize mucin-domain glycoproteins from cell lysate and patient biofluids.
- Stacy A. Malaker
- , Nicholas M. Riley
- & Carolyn R. Bertozzi
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Article
| Open AccessMechanism of glycogen synthase inactivation and interaction with glycogenin
Glycogen is a major energy reserve in eukaryotes and is synthesised in part by glycogenin (GN) and glycogen synthase (GS). Here, authors describe the structural basis of GS regulation, specifically the mechanism of inactivation by phosphorylation.
- Laura Marr
- , Dipsikha Biswas
- & Elton Zeqiraj
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Article
| Open AccessThe structure of EXTL3 helps to explain the different roles of bi-domain exostosins in heparan sulfate synthesis
Heparan sulphate (HS)—a common cell surface decoration—is a carbohydrate of alternating sugars assembled by bi-domain enzymes such as EXTL3. Here, authors present the structure of EXTL3, explain EXTL3 lost activity, and propose that HS extension is distributive.
- L. F. L. Wilson
- , T. Dendooven
- & P. Dupree
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Article
| Open AccessMembrane curvature regulates the spatial distribution of bulky glycoproteins
MUC1 is a heavily glycosylated protein on the cell surface. Here the authors show that MUC1 prefers negative over positive membrane curvature due to its bulky size, enabling MUC1 to avoid endocytosis and surface removal based on curvature preference.
- Chih-Hao Lu
- , Kayvon Pedram
- & Bianxiao Cui
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Article
| Open AccessO-GlcNAc modification of leucyl-tRNA synthetase 1 integrates leucine and glucose availability to regulate mTORC1 and the metabolic fate of leucine
Leucyl-tRNA synthetase 1 (LARS1) is a leucine sensor for mTORC1 signaling and regulates leucine utilization depending on glucose availability. Here, the author show that O-GlcNAcylation of LARS1 is crucial for its ability to regulate mTORC1 activity and leucine metabolism upon glucose starvation.
- Kibum Kim
- , Hee Chan Yoo
- & Jung Min Han
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Article
| Open AccessOrigins of glycan selectivity in streptococcal Siglec-like adhesins suggest mechanisms of receptor adaptation
Streptococcal siglec-like binding regions (SLBRs) selectively bind cell surface glycans, but the basis for this selectivity is not understood. Here, the authors identify selectivity-modulating SLBR regions and study how changes in SLBR glycan selectivity affect interactions with human glycoprotein receptors.
- Barbara A. Bensing
- , Haley E. Stubbs
- & T. M. Iverson
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Article
| Open AccessCa2+-mediated higher-order assembly of heterodimers in amino acid transport system b0,+ biogenesis and cystinuria
Cystinuria is caused by mutations in heterodimeric amino acid transporter known as system b0,+. Here, authors discover that Ca2+ stabilizes the interface between two system b0,+ regulatory subunits rBAT, leading to super-dimerization of the b0,+AT–rBAT heterodimer, facilitating system b0,+ maturation.
- Yongchan Lee
- , Pattama Wiriyasermkul
- & Shushi Nagamori
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Article
| Open AccessStructural basis for the synthesis of the core 1 structure by C1GalT1
The glycosyltransferase C1GalT1 directs a key step in protein O-glycosylation important for the expression of the cancer-associated Tn and T antigens. Here, the authors provide molecular insights into the function of C1GalT1 by solving the crystal structure of the Drosophila enzyme-substrate complex.
- Andrés Manuel González-Ramírez
- , Ana Sofia Grosso
- & Ramon Hurtado-Guerrero
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Article
| Open AccessCDP-ribitol prodrug treatment ameliorates ISPD-deficient muscular dystrophy mouse model
Ribitol-phospate modification is essential for the function of α-dystroglycan, and mutations in ISPD, an enzyme that synthesizes the the ribitol-phosphate donor CDP-ribitol, cause muscular dystrophy. Here, the authors show that recovery of CDP-ribitol levels, either via AAV-mediated gene therapy or prodrug treatment, rescues dystroglycan function and pathology in a mouse model.
- Hideki Tokuoka
- , Rieko Imae
- & Tatsushi Toda
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Article
| Open AccessMechanism of cooperative N-glycan processing by the multi-modular endoglycosidase EndoE
EndoE is a multi-domain glycoside hydrolase of the human pathogen Enterococcus faecalis. Here, the authors present crystal structures of EndoE and provide biochemical insights into the molecular basis of EndoE’s substrate specificity and catalytic mechanism.
- Mikel García-Alija
- , Jonathan J. Du
- & Marcelo E. Guerin
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| Open AccessMammalian brain glycoproteins exhibit diminished glycan complexity compared to other tissues
Protein glycosylation is critical in brain development and disease. Here, the authors characterize brain glycans in detail, showing that they are simpler and more homogenous than glycans from other tissues and providing a basis for future studies of brain glycosylation.
- Sarah E. Williams
- , Maxence Noel
- & Robert G. Mealer
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Article
| Open AccessTransglutaminase 3 crosslinks the secreted gel-forming mucus component Mucin-2 and stabilizes the colonic mucus layer
The colonic mucus layer is an organized system providing a physical barrier against pathogens and simultaneously harbouring the commensal flora. Here the authors report that transglutaminase 3 activity contributes to homeostasis of the colonic mucus layer and the lack of this enzymatic activity leads to increased susceptibility against DSS-induced colitis in mice.
- Jack D. A. Sharpen
- , Brendan Dolan
- & Christian V. Recktenwald
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Article
| Open AccessNeutron crystallography reveals mechanisms used by Pseudomonas aeruginosa for host-cell binding
Pseudomonas aeruginosa employs lectins to bind to its host cells, and is known to be the major cause of lung infections. Lectin B (LecB) from Pseudomonas aeruginosa binds specifically to galactose and fucose and is important for pathogenicity, adhesion and biofilm formation. In this work, the neutron crystal structure (1.9 Å) of the deuterated LecB/Ca/fucose complex is reported. The structure, in combination with perdeuteration of the ligand and the receptor allowed the observation of hydrogen atoms, protonation states and hydrogen bonds involved in the interaction between pathogenic bacteria and host cells. Thus the study provides structural insights into the mechanism of high affinity binding of LecB to its targets.
- Lukas Gajdos
- , Matthew P. Blakeley
- & Anne Imberty
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Article
| Open AccessDiscovery of indole-modified aptamers for highly specific recognition of protein glycoforms
Glycosylation is an abundant form of post-translational modification. Here the authors present a generalizable workflow for the selection of indole-modified aptamers that can recognize protein glycoforms with high specificity.
- Alex M. Yoshikawa
- , Alexandra Rangel
- & H. Tom Soh
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Article
| Open AccessCatalytic flexibility of rice glycosyltransferase OsUGT91C1 for the production of palatable steviol glycosides
Steviol glycosides from the plant Stevia rebaudiana are already used as lowcalorie sweeteners, but the most abundant naturally occurring compounds have a bitter aftertaste. Here, the authors characterize and engineer rice glycosyltransferase OsUGT91C1 to facilitate the large-scale production of naturally rare but palatable glycosides Reb D and Reb M
- Jinzhu Zhang
- , Minghai Tang
- & Wei Cheng
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Article
| Open AccessCryo-EM structure provides insights into the dimer arrangement of the O-linked β-N-acetylglucosamine transferase OGT
The modification of proteins with O-linked β-N-acetylglucosamine (OGlcNAc) plays roles in regulation of numerous cellular functions while incorrect O-GlcNAcylation patterns are linked to disease. Here, the authors report a cryo-EM structure of full-length O-GlcNAc transferase (OGT), the only enzyme responsible for O-GlcNAcylation.
- Richard W. Meek
- , James N. Blaza
- & Gideon J. Davies
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Article
| Open AccessCongenital disorder of glycosylation caused by starting site-specific variant in syntaxin-5
Mutations in genes critical for proper intra-Golgi transport can cause human syndromes due to defects in glycosylation of proteins. Here, the authors identify a human variant of Syntaxin-5 that causes fatal multisystem disease and mislocalization of glycosyltransferases due to altered Golgi transport.
- Peter T. A. Linders
- , Eveline C. F. Gerretsen
- & Geert van den Bogaart
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Article
| Open AccessGproDIA enables data-independent acquisition glycoproteomics with comprehensive statistical control
Data independent acquisition (DIA) proteomics provides deep coverage and high quantitative accuracy, but is not yet well established in glycoproteomics. Here, the authors develop a DIA-based glycoproteomics workflow with stringent statistical controls to enable accurate glycopeptide identification.
- Yi Yang
- , Guoquan Yan
- & Liang Qiao
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Article
| Open AccessAfucosylated Plasmodium falciparum-specific IgG is induced by infection but not by subunit vaccination
Here, Larsen et al. describe differences in Fc fucosylation of P. falciparum PfEMP1-specific IgG produced in response to natural infection versus VAR2CSA-type subunit vaccination, which leads to differences in the ability to induce FcγRIIIa-dependent natural killer cell degranulation.
- Mads Delbo Larsen
- , Mary Lopez-Perez
- & Gestur Vidarsson
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Article
| Open AccessGlycan remodeled erythrocytes facilitate antigenic characterization of recent A/H3N2 influenza viruses
Here, Broszeit et al. show that circulating A/H3N2 viruses have evolved binding specificity to α2,6-sialosides on extended LacNAc moieties and therefore cannot agglutinate erythrocytes. Applying glycan remodeling allows to install functional receptors on erythrocytes and promotes identification of newly circulating variants to facilitate vaccine design.
- Frederik Broszeit
- , Rosanne J. van Beek
- & Geert-Jan Boons
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Article
| Open AccessCorrecting for sparsity and interdependence in glycomics by accounting for glycan biosynthesis
Glycomics can uncover important molecular changes but measured glycans are highly interconnected and incompatible with common statistical methods, introducing pitfalls during analysis. Here, the authors develop an approach to identify glycan dependencies across samples to facilitate comparative glycomics.
- Bokan Bao
- , Benjamin P. Kellman
- & Nathan E. Lewis
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Article
| Open AccessDisplay of the human mucinome with defined O-glycans by gene engineered cells
Mucins play critical roles in maintaining the human microbiome, with their O-glycosylated tandem repeats (TRs) providing important cues for microbiota. Here, the authors develop a cellular platform for producing TRs with defined O-glycan structures to dissect the functions of TR O-glycosylation.
- Rebecca Nason
- , Christian Büll
- & Yoshiki Narimatsu
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Article
| Open AccessXyloglucan processing machinery in Xanthomonas pathogens and its role in the transcriptional activation of virulence factors
Xyloglucans are polysaccharides found in plant cell walls. Here, the authors describe the xyloglucan depolymerization machinery of phytopathogenic Xanthomonas bacteria, and show that sugars released by this system induce the expression of key virulence factors driving pathogenesis.
- Plinio S. Vieira
- , Isabela M. Bonfim
- & Mario T. Murakami
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| Open AccessChemoenzymatic modular assembly of O-GalNAc glycans for functional glycomics
O-GalNAc glycans are essential in many biological and pathological processes, but difficult to access due to their structural complexity and synthetic challenges. Here, the authors report an efficient chemoenzymatic modular assembly strategy to construct structurally diverse O-GalNAc glycans, use the synthesised glycans to generate a synthetic mucin O-glycan microarray and profile binding specificities of glycan-binding proteins.
- Shuaishuai Wang
- , Congcong Chen
- & Lei Li
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Article
| Open AccessSynergistic sequence contributions bias glycation outcomes
Advanced glycation end-products (AGEs), such as methylglyoxal-derived hydroimidazolone isomer (MGH-1), are associated with disease and age-related disorders, and occur spontaneously, so it is unclear why specific protein sites become modified with specific AGEs. Here, the authors use a combinatorial peptide library to determine the chemical features that favour MGH-1 formation for short peptides and demonstrate a key role of tyrosine in this process.
- Joseph M. McEwen
- , Sasha Fraser
- & Rebecca A. Scheck
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Article
| Open AccessFKRP-dependent glycosylation of fibronectin regulates muscle pathology in muscular dystrophy
FKRP mutations cause muscular dystrophies with varied clinical presentations. The target of FKRP is α-dystroglycan, but here the authors show that FKRP also directs sialylation of fibronectin, a process that is essential for recruitment o collagen to the muscle basement membrane.
- A. J. Wood
- , C. H. Lin
- & P. D. Currie
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Article
| Open AccessLoss of α2-6 sialylation promotes the transformation of synovial fibroblasts into a pro-inflammatory phenotype in arthritis
Dysregulation of synovial fibroblasts is thought to be an important step in the pathogenesis of rheumatoid arthritis. Here the authors implicate α2-6 sialylation in this process by studying the glycome of these cells in patients and in a mouse model of inflammatory joint disease.
- Yilin Wang
- , Aneesah Khan
- & Miguel A. Pineda
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Article
| Open AccessProtein kinase A controls the hexosamine pathway by tuning the feedback inhibition of GFAT-1
The glutamine fructose-6-phosphate amidotransferase 1 (GFAT-1) is the rate-limiting enzyme in the hexosamine pathway producing uridine 5’-diphospho-N-acetyl-D-glucosamine (UDP-GlcNAc), an essential glycosylation precursor. Here, the authors dissect the mechanisms of GFAT-1 regulation by protein kinase A (PKA)-mediated phosphorylation.
- Sabine Ruegenberg
- , Felix A. M. C. Mayr
- & Martin S. Denzel
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Article
| Open AccessRed blood cell mannoses as phagocytic ligands mediating both sickle cell anaemia and malaria resistance
Red blood cells (RBCs) are phagocytosed in the spleen in sickle cell disease and malaria. Here, Cao et al. show that high mannose N-glycans, exposed on diseased or oxidized RBC surfaces, bind mannose receptor CD206 on host cells, mediating phagocytosis.
- Huan Cao
- , Aristotelis Antonopoulos
- & Mark A. Vickers
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Article
| Open AccessGlycan chip based on structure-switchable DNA linker for on-chip biosynthesis of cancer-associated complex glycans
Current methods for on-chip glycan biosynthesis suffer from analysing products, often resulting in poor purity and yield. Here the authors report a glycan chip developed by introducing a pH-responsive i-motif DNA linker to control the immobilization and isolation of glycans on chips.
- Hye Ryoung Heo
- , Kye Il Joo
- & Hyung Joon Cha
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Article
| Open AccessSialic acids in pancreatic cancer cells drive tumour-associated macrophage differentiation via the Siglec receptors Siglec-7 and Siglec-9
Alterations in glycosylation in tumours facilitate tumour progression. Here, the authors show that pancreatic ductal adenocarcinomas present increased sialylation, which stimulates the polarisation of monocytes via Siglec receptors, resulting in the generation of immune suppressive tumour associated macrophages.
- Ernesto Rodriguez
- , Kelly Boelaars
- & Yvette van Kooyk
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Article
| Open AccessUnravelling the structural complexity of glycolipids with cryogenic infrared spectroscopy
Glycolipids are glycoconjugates with important biological functions, but techniques for their analysis are deficient. Here, the authors report the use of cryogenic gas-phase infrared spectroscopy to investigate isomerism in a set of immunologically relevant glycolipids, and show that their structural features can be accurately resolved based on a narrow spectral fingerprint region.
- Carla Kirschbaum
- , Kim Greis
- & Kevin Pagel
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Article
| Open AccessDiatom fucan polysaccharide precipitates carbon during algal blooms
The fate of ocean carbon is determined by the balance between primary productivity and heterotrophic breakdown of that photosynthate. Here the authors show that diatoms produce a polysaccharide that resists bacterial degradation, accumulates, aggregates and stores carbon during spring blooms.
- Silvia Vidal-Melgosa
- , Andreas Sichert
- & Jan-Hendrik Hehemann
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Article
| Open AccessTwo distinct catalytic pathways for GH43 xylanolytic enzymes unveiled by X-ray and QM/MM simulations
Family 43 glycoside hydrolases (GH43) are involved in the breakdown of hemicellulose. Functional, structural and computational characterization of a GH43 enzyme, including a snapshot of an active Michaelis complex, reveal the hydrolysis mechanism and suggest two possible reaction pathways.
- Mariana A. B. Morais
- , Joan Coines
- & Mario T. Murakami
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Article
| Open AccessAssembling custom side chains on proteoglycans to interrogate their function in living cells
Currently, it is not possible to generate proteoglycans displaying glycosaminoglycan chains with specific structures. Here the authors show that by using an aldehyde tag-based methodology it is possible to insert these specific chains onto proteoglycans expressed on the cell surface.
- Wenshuang Wang
- , Naihan Han
- & Fuchuan Li
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Article
| Open AccessProteomic profiling and genome-wide mapping of O-GlcNAc chromatin-associated proteins reveal an O-GlcNAc-regulated genotoxic stress response
Protein O-GlcNAcylation is involved in regulating gene expression and maintaining cellular homeostasis. Here, the authors develop a chemical reporter-based strategy for the proteomic profiling and genome-wide mapping of genotoxic stress-induced O-GlcNAcylated chromatin-associated proteins.
- Yubo Liu
- , Qiushi Chen
- & Jianing Zhang
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Article
| Open AccessA streamlined pipeline for multiplexed quantitative site-specific N-glycoproteomics
Comprehensive quantitative profiling of intact glycopeptides remains technically challenging. To address this, the authors here develop an integrated quantitative glycoproteomic workflow, including optimized sample preparation, multiplexed quantification and a dedicated data processing tool.
- Pan Fang
- , Yanlong Ji
- & Henning Urlaub
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Article
| Open AccessExploring and applying the substrate promiscuity of a C-glycosyltransferase in the chemo-enzymatic synthesis of bioactive C-glycosides
C-glycosides are of pharmaceutical interest due to their stability against in vivo hydrolysis, however their enzymatic synthesis faces challenges. Here, the authors report a C-glycosyltransferase from Aloe barbadensis catalysing the C-glycosylation of drug-like acceptors to generate bioactive C-glycosides.
- Kebo Xie
- , Xiaolin Zhang
- & Jungui Dai
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Article
| Open AccessA versatile soluble siglec scaffold for sensitive and quantitative detection of glycan ligands
Sialic acid-binding immunoglobulin-type lectins (Siglecs) are a family of immunomodulatory receptors expressed on cells of the hematopoietic lineage. Here the authors demonstrate an approach for the identification of the glycan ligands of Siglecs, which is also applicable to other families of glycan-binding proteins.
- Emily Rodrigues
- , Jaesoo Jung
- & Matthew S. Macauley
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Article
| Open AccessThioglycoligase derived from fungal GH3 β-xylosidase is a multi-glycoligase with broad acceptor tolerance
Thioglycoligases have proved useful for bonding carbohydrates to non-sugar acceptors, however, the scope of these biocatalysts is usually limited. Here, the authors engineer a xylosidase into a thioglycoligase with the ability to form O-, N-, S- and Se- glycosides together with sugar esters and phosphoesters.
- Manuel Nieto-Domínguez
- , Beatriz Fernández de Toro
- & María Jesús Martínez
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Article
| Open AccessStructural basis of mammalian mucin processing by the human gut O-glycopeptidase OgpA from Akkermansia muciniphila
OgpA is an O-glycopeptidase from Akkermansia muciniphila, a mucin-degrading bacterium commonly found in the human gut. A thorough characterization of OgpA, including crystal structures in complex with substrate or product, reveals molecular basis of O-glycan recognition and enzyme specificity.
- Beatriz Trastoy
- , Andreas Naegeli
- & Marcelo E. Guerin
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Article
| Open AccessChemical synthesis of glycans up to a 128-mer relevant to the O-antigen of Bacteroides vulgatus
Glycans are abundant biomolecules that mediate essential biological processes, but their chemical synthesis is challenging. Here, the authors report the synthesis of glycans up to a 128-mer, which represents the O-antigen of Bacteroides vulgatus lipopolysaccharide and one of the longest synthetic glycans to date.
- Qian Zhu
- , Zhengnan Shen
- & Biao Yu