Glycobiology最新文献

Human lectins are critical carbohydrate-binding proteins that recognize diverse glycoconjugates from microorganisms and can play a key role in host-microbe interactions. Despite their importance in immune recognition and microbe binding, the specific glycan ligands and functions of many human lectins remain poorly understood. Using previous proof-of-concept studies on selected lectins as the foundation for this work, we present ten additional glycan analysis probes (GAPs) from a diverse set of human soluble lectins, offering robust tools to investigate glycan-mediated interactions. We describe a protein engineering platform that enables scalable production of GAPs that maintain native-like conformations and oligomerization states, equipped with functional reporter tags for targeted glycan profiling. We demonstrate that the soluble GAP reagents can be used in various applications, including glycan array analysis, mucin-binding assays, tissue staining, and microbe binding in complex populations. These capabilities make GAPs valuable for dissecting interactions relevant to understanding host responses to microbes. The tools can also be used to probe differential microbial and mammalian glycan interactions, which are crucial for understanding the interactions of lectins in a physiological environment where both glycan types exist. GAPs have potential as diagnostic and prognostic tools for detecting glycan alterations in chronic diseases, microbial dysbiosis, and immune-related conditions.

Galectins are a family of β-galactosides-binding protein, crucial regulators of host-virus interactions. They achieve this by recognizing specific glycan patterns on viral surfaces or mediating interactions with intracellular viral or host proteins, subsequently influencing the critical phases of the viral life cycle, such as attachment, replication, immune evasion, and reactivation. Furthermore, galectins modulate host immune responses, shaping the progression and outcomes of viral infections. This review comprehensively examines the roles of both endogenous and exogenous galectins in viral infections, noting that only a few galectins, including Galectin-1, -3, -4, -7, -8, and -9, Have been identified as key players in viral infection. Notably, Galectin-1, -3, and -9 play diverse functions in both DNA and RNA viral infection. Emerging evidence highlights the potential of Galectin-4 and -8 as intracellular sensors and modulators of viral pathogenesis. Endogenous galectins, produced by host cells, act through both glycan-dependent and glycan-independent mechanisms, influencing viral processes and immune responses. Exogenous galectins, which are secreted by other cells or administered as recombinant proteins, can either enhance or counteract the actions of endogenous galectins. The functions of galectins are virus-specific and context-dependent, serving as either promoters or inhibitors of viral replication and reactivation. Dysregulation of galectin expression is often linked to disease progression, highlighting their potential as diagnostic and prognostic biomarkers, as well as therapeutic targets. The important and varied roles that galectins play in viral infections are highlighted in this review, which also provides fresh insights into host-pathogen interactions and the development of antiviral tactics.

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Coley's Toxin comprised a mixture of cell-free, heat-treated culture media from Streptococcus pyogenes (originally Streptococus erysipelatos) and Serratia marcescens (originally Bacillus prodigiosus). A 250 kDa tumor hemorrhage-inducing polysaccharide "PS1" is reported here secreted into culture medium by S. marcescens. Four h after PS1 is injected at 32 μg/kg (10pM) into the tail vein of Balb/C mice bearing C26 subcutaneous colon-derived tumors, tumor-specific capillary hemorrhage is exhibited in 90% of tumors. As a positive control, CM101, a similar tumor hemorrhagic polysaccharide from Streptococcus agalactica caused tumor hemorrhage in 75% of tumors in the Balb/C-C26 model at 7.5 μg/kg(2.5pM). CM101 has previously been safety tested in a Phase I clinical trial. These two polysaccharides have merit to be identified as the active principal ingredients (API's) of Coley'sToxin. Additional approaches to cancer therapy are a global need. No matter the level of wealth of victims, some cancers are still incurable. Recall in recent years the tragic early cancer deaths of Steve Jobs and Paul Allen among other luminaries. Streptococcal and Serratia bacterial extracts have unique tumor specific capillary destructive activity, with observations originating with sarcomas cured by nosocomial  erysipelas  infections in the 1860's. The active pharmaceutical ingredients (API's) in these extracts and Coley's Toxins are proposed to be polysaccharides.

N-glycosylation is a critical quality attribute to consider when expressing recombinant glycoproteins in eukaryotic cells including plant cells. N-acetylglucosaminyltransferase I (GnTI) initiates complex N-glycan maturation in the Golgi apparatus by transferring a single N-acetylglucosamine (GlcNAc) residue on the alpha1,3-arm of a Man5 N-glycan acceptor. This step is required for the processing of high mannose into hybrid and complex N-glycans. Therefore, Arabidopsis mutants lacking GnTI activity display accumulation of Man5 N-glycans instead of complex N-glycans and do not synthesise N-glycans containing core alpha1,3-fucose residue. In contrast, GnTI knockout cell line of Nicotiana tabacum BY-2 still displays a little core alpha1,3-fucose signal on western blotting. Here, we show that N. tabacum contains two alpha1,3-fucosyltransferase types, one of which is able to transfer a core alpha1,3-fucose on a Man5 substrate when no Man5Gn substrate is available such as in BY-2 GnTI knock-out cell lines.

The ephrin family of membrane proteins mediate intracellular signalling as ligands of transmembrane Eph tyrosine kinase receptors during cell-cell interactions. Ephrin/Eph signalling regulates processes like cell migration and angiogenesis and is of particular importance during embryonic development. Ephrins-A3 and -B3 can also bind to cell surface-associated and soluble heparan sulfate proteoglycans (HSPGs) that also play important roles during early development. Here we show that ephrins-B1, -B2, and -B3 all can bind in cis to cell surface HSPGs, while only ephrin-B1 interacts with cell surface HSPGs in a way that retards HSPG endocytosis. Expressing ephrin-B1 in HEK293T cells, using polyethyleneimine (PEI) as transfection agent, increased cell surface levels of HSPGs which were detected by an anti-heparan sulfate (HS) antibody or by ephrin-B3-Fc binding. Ephrin-B1 in the plasma membrane seemed to retard PEI-induced HSPG internalisation and degradation. Binding of HSPGs by ephrin-B1 was observed for the human, mouse, xenopus, and zebrafish homologs, and did not require the cytoplasmic tail of ephrin-B1 that contains tyrosines shown to be involved in intracellular signalling. Furthermore, ephrin-B1 could bind the HSPG variant of CD44 (CD44V3-10), a complex that could further associate with fibroblast growth factor receptors (1 and 4) after co-expression with one of these receptors. In summary, our data indicate that ephrin-B1 can regulate cellular HSPG turnover and is able to form complexes of potential biological importance with CD44V3-10 and fibroblast growth factor receptors.

Mucin-type O-linked glycosylation is initiated by the transfer of a single N-acetyl-D-galactosamine (GalNAc) to the hydroxyl group of either a serine (Ser) or threonine (Thr) residue. This process is catalysed by a portfolio of twenty isoenzymes, the UDP-N-acetyl-α-D-galactosamine:polypeptide N-acetylgalactosaminyltransferases (ppGalNAc-Ts, GalNAc-Ts or GALNTs) to create the Thomsen nouvelle (Tn) antigen (GalNAcα1-O-Ser/Thr ). In healthy adult cells, Tn antigen is further elaborated by the action of specific glycosyltransferases to either form one of eight core structures, which themselves can be extended to form more complex glycans, or into sialyl Tn or sialyl core 1 (sialyl T), where sialylation terminates chain extension. These O-glycans, produced through mucin-type O-linked glycosylation, are a feature of many secreted and membrane-bound proteins, and are fundamental in a wide range of biological functions. Dysregulation of this process, often resulting in the exposure of usually cryptic truncated O-glycans including Tn antigen, is important in a wide range of pathologies and has been implicated in cancer metastasis. The regulation of mucin-type O-linked glycosylation, in health and disease, is highly complex and not fully understood. It is determined by a myriad of mechanisms, from transcriptional control, mutation, posttranslational control, stability of transferases, their relocation within the secretory pathway, and changes in the fundamental structure and environment of the Golgi apparatus. This review presents an overview of the evidence for these potential regulatory steps in the synthesis of truncated mucin-type O-linked glycans in cancer.

Heparan-6-O-endosulfatase 2 (Sulf-2) is a proteoglycan enzyme that modifies sulfation of heparan sulfate proteoglycans. Dysregulation of Sulf-2 is associated with various pathological conditions, including cancer, which makes Sulf-2 a potential therapeutic target. Despite the key pathophysiological roles of Sulf-2, inhibitors remain insufficiently developed. In previous work, a fucosylated chondroitin sulfate from the sea cucumber Holothuria floridana (HfFucCS) exhibited potent Sulf-2 inhibition. This study investigates the structural basis of HfFucCS-mediated Sulf-2 inhibition, examines the binding profile of HfFucCS to Sulf-2, and explores the mode of inhibition. Additionally, a structurally diverse library of sulfated poly/oligosaccharides, including common glycosaminoglycans and unique marine sulfated glycans, was screened for Sulf-2 inhibition. Results from a high-throughput arylsulfatase assay and specific 6-O-desulfation assay have proved that HfFucCS is the most potent among the tested sulfated glycans, likely due to the presence of the unique 3,4-disulfated fucose structural motif. HfFucCS demonstrated non-competitive inhibition, and inhibitory analysis of its low-molecular-weight fragments suggests a minimum length of ~7.5 kDa for effective inhibition. Surface plasmon resonance analyses revealed that Sulf-2 binds to surface heparin with high affinity (KD of 0.817 nM). HfFucCS and its derivatives effectively disrupt this interaction. Results from mass spectrometry-hydroxyl radical protein footprinting and repulsive scaling replica exchange molecular dynamics indicate similarities in the binding of heparin and HfFucCS oligosaccharides to both the catalytic and hydrophilic domains of Sulf-2. These findings reveal the unique inhibitory properties of a structurally distinct marine glycosaminoglycan, supporting its further investigation as a selective and effective inhibitor for Sulf-2-associated cancer events.

Glycosphingolipids are a unique class of bioactive lipids responsible for lateral membrane organization and signaling found in high abundance in the central nervous system. Using nanoflow MEA Chip Q/ToF mass spectrometry, we profiled the intact glycosphingolipids of the elderly human brain in a region-specific manner. By chromatographic separation of glycan and ceramide isomers, we determined gangliosides to be the highest source of heterogeneity between regions with the expression of a- and b-series glycan structures. Investigation of these trends showed that specific glycan structures were, in part, determined by the structure of their lipid backbone. This study provides insight into the dynamic process of membrane remodeling in the brain during aging.