The initial contact of pathogens with host cells is usually mediated by their adhesion to glycan structures present on the cell surface in order to enable infection. Furthermore, glycans play ...important roles in the modulation of the host immune responses to infection. Understanding the carbohydrate-pathogen interactions are of importance for the development of novel and efficient strategies to either prevent, or interfere with pathogenic infection. Synthetic glycopeptides and mimetics thereof are capable of imitating the multivalent display of carbohydrates at the cell surface, which have become an important objective of research over the last decade. Glycopeptide based constructs may function as vaccines or anti-adhesive agents that interfere with the ability of pathogens to adhere to the host cell glycans and thus possess the potential to improve or replace treatments that suffer from resistance. Additionally, synthetic glycopeptides are used as tools for epitope mapping of antibodies directed against structures present on various pathogens and have become important to improve serodiagnostic methods and to develop novel epitope-based vaccines. This review will provide an overview of the most recent advances in the synthesis and application of glycopeptides and glycopeptide mimetics exhibiting a peptide-like backbone in glycobiology.
Multivalent interactions at biological interfaces occur frequently in nature and mediate recognition and interactions in essential physiological processes such as cell-to-cell adhesion. Multivalency ...is also a key principle that allows tight binding between pathogens and host cells during the initial stages of infection. One promising approach to prevent infection is the design of synthetic or semisynthetic multivalent binders that interfere with pathogen adhesion
. Here, we present a multivalent binder that is based on a spatially defined arrangement of ligands for the viral spike protein haemagglutinin of the influenza A virus. Complementary experimental and theoretical approaches demonstrate that bacteriophage capsids, which carry host cell haemagglutinin ligands in an arrangement matching the geometry of binding sites of the spike protein, can bind to viruses in a defined multivalent mode. These capsids cover the entire virus envelope, thus preventing its binding to the host cell as visualized by cryo-electron tomography. As a consequence, virus infection can be inhibited in vitro, ex vivo and in vivo. Such highly functionalized capsids present an alternative to strategies that target virus entry by spike-inhibiting antibodies
and peptides
or that address late steps of the viral replication cycle
.
Photochemical degradation is an important issue to be overcome in advancing the lifetime of fluorene-containing conjugated polymers. In order to optimize the inertness of the materials, a ...quantitative measure for the efficiency of degradation is needed. Here, we introduce a method to measure a relative quantum yield of the photochemical degradation by monitoring the kinetics of the process by means of UV/vis spectroscopy and liquid chromatography (LC) techniques. This method is employed to a set of differently substituted 2,7-diphenylfluorenes, serving as model compounds for polyfluorene materials. Our measurements show that the quantum yield changes by orders of magnitude upon varying the bridge substituents and that altered kinetics indicate changing degradation mechanisms.
The interactions between bacterial lectins and carbohydrates on the host cell surface can mediate bacterial adhesion, invasion, and immune evasion. Multivalency plays a key role in these binding ...events. However, additional molecular mechanisms greatly impact multivalent binding recognition. To develop specific and effective bacterial inhibitors, a deeper understanding of the complex underlying mechanisms of bacterial adhesion processes is necessary. By interfering with bacterial adhesion, synthetic multivalent glycoconjugates do not only have the potential to improve or replace antibiotic treatments, but also represent useful tools to study carbohydrate‐pathogen interactions. In this review, we highlight a few recent advances in the synthesis and application of synthetic glycan‐based scaffolds to uncover the nature of glycan‐bacteria interactions and to design efficient bacterial inhibitors.
Sweet and sticky: This review describes recent advances in the synthesis and application of homo‐and heterovalent glycan‐scaffolds as potential candidates for efficient bacteria lectin inhibition. To study and address the many complex molecular mechanisms involved in multivalent lectin binding events, novel glycoconjugate designs and fine‐tunable glycomaterials have been developed.
In the last decade, it was discovered that protein mucin‐type O‐glycosylation and O‐GlcNAcylation modify Tyr residues besides the well explored Thr and Ser amino acids. Several glycoproteomic studies ...have identified α‐GalNAc‐O‐Tyr modifications, and studies propose that β‐GlcNAc‐O‐Tyr also exists as a new group of posttranslational modifications (PTMs). Specific bacterial toxins have further been identified to modify host GTPases with α‐GlcNAc‐O‐Tyr to promote bacterial virulence. Despite being identified on numerous proteins, the biological roles, biosynthesis and expression of GalNAc‐ and GlcNAc‐O‐Tyr modifications are poorly understood. A major obstacle is the lack of tools to specifically detect and identify proteins containing these modifications. With this in mind, we prepared vaccine constructs and raised antibodies to enable selective detection of proteins carrying these new PTMs. The obtained polyclonal antibody sera were evaluated using ELISA and glycopeptide microarrays and were found to be highly selective for GlcNAc‐ and GalNAc‐O‐Tyr glycopeptides over the corresponding Ser‐ and Thr‐modifications. For microarray analysis, synthetic GlcNAc‐ and GalNAc‐O‐Tyr Fmoc‐amino acids were prepared and applied in Fmoc‐SPPS to obtain an extensive O‐glycopeptide library. After affinity purification, the antibodies were applied in western blot analysis and showed specific detection of α‐GlcNAc‐O‐Tyr modified RhoA GTPase.
Catch as catch can. Fmoc‐SPPS α‐GalNAc‐, β‐GlcNAc‐ and α‐GlcNAc‐O‐Tyr amino acid building blocks were prepared and applied in synthesis of glycopeptide vaccine constructs and an extensive Tyr‐modified glycopeptide library. Antibodies were raised and affinity purified against the antigen peptides. Evaluation by ELISA, glycopeptide microarrays and by western blot analysis showed high antibody specificity towards GalNAc‐ and GlcNAc‐O‐Tyr modifications without cross reactivity to the corresponding Ser or Thr glycopeptides.
Mucin glycoproteins are essential components of the mucosal barrier, which protects the host from pathogens. Throughout evolution, bacteria have developed strategies to modulate and penetrate this ...barrier, and cause virulence by interacting with mucin O‐glycans at the epithelial cell‐surface. O‐fucosylated glycan epitopes on mucins are key ligands of many bacterial lectins. Here, a chemoenzymatic synthesis strategy is described to prepare a library of fucosylated mucin core glycopeptides to enable studies of mucin‐interacting and fucose‐binding bacterial lectins. Glycan cores with biologically important Lewis and H‐antigens were prepared decorating the peptide backbone at different sites and densities. The fucosylated mucin glycopeptides were applied in microarray binding studies to explore the importance of glycan core and peptide backbone presentation of these antigens in binding interactions with the P. aeruginosa lectin LecB and the C. difficile toxin A.
To enable studies of fucose‐binding bacterial lectins, a library of fucosylated mucin glycopeptides carrying Lewis‐ and H‐antigen structures on mucin core glycans was prepared using a chemoenzymatic approach. This library was applied to evaluate the Pseudomonas aeruginosa lectin LecB and the Clostridium difficile toxin A binding preferences to explore the importance of the glycan core and peptide backbone presentation for lectin recognition.
O‐fucosylated glycan epitopes of membrane‐bound mucin glycoproteins are key ligands of many bacterial lectins. In their Research Article (e202302437), Ulrika Westerlind and co‐workers describe the ...synthesis of fucosylated mucin glycopeptides using a chemoenzymatic approach. Starting from simple building blocks, Fmoc‐protected glycosylated amino acids were prepared and incorporated into peptides and enzymatically diversified. Bacteria lectin interactions were then elucidated on glycopeptide microarrays.
MUC1 glycopeptides are attractive antigens for anti-cancer vaccine development. One potential drawback in using the native MUC1 glycopeptide for vaccine design is the instability of the
O
-glycosyl ...linkage between the glycan and the peptide backbone to glycosidase. To overcome this challenge, a MUC1 glycopeptide mimic has been synthesized with the galactose-galactosamine disaccharide linked with threonine (Thomsen-Friedenreich or Tf antigen) through an unnatural β-glycosyl bond. The resulting MUC1-β-Tf had a much-enhanced stability toward a glycosidase capable of cleaving the glycan from the corresponding MUC1 glycopeptide with the natural α-Tf linkage. The MUC1-β-Tf was subsequently conjugated with a powerful carrier bacteriophage Qβ. The conjugate induced high levels of IgG antibodies in clinically relevant human MUC1 transgenic mice, which cross-recognized not only the natural MUC1-α-Tf glycopeptide but also MUC1 expressing tumor cells, supporting the notion that a simple switch of the stereochemistry of the glycan/peptide linkage can be a strategy for anti-cancer vaccine epitope design for glycopeptides.
A MUC1 glycopeptide bearing an unnatural β-glycosyl bond between the glycan and the peptide backbone was synthesized. The mimic can induce high levels of IgG antibodies cross-recognizing cancer cells expressing the native MUC1 glycoprotein.
Mucin expression and glycosylation patterns on cancer cells differ markedly from healthy cells. Mucin 1 (MUC1) is overexpressed in several solid tumors and presents high levels of aberrant, truncated ...O-glycans (e.g., Tn antigen). Dendritic cells (DCs) express lectins that bind to these tumor-associated carbohydrate antigens (TACAs) to modulate immune responses. Selectively targeting these receptors with synthetic TACAs is a promising strategy to develop anticancer vaccines and to overcome TACA tolerance. In this work, we prepared, via a solid phase peptide synthesis approach, a modular tripartite vaccine candidate, incorporating a high-affinity glycocluster based on a tetraphenylethylene scaffold, to target the macrophage galactose-type lectin (MGL) on antigen presenting cells. MGL is a C-type lectin receptor that binds Tn antigens and can route them to human leukocyte antigen class II or I, making it an attractive target for anticancer vaccines. Conjugation of the glycocluster to a library of MUC1 glycopeptides bearing the Tn antigen is shown to promote uptake and recognition of the TACA by DCs via MGL. In vivo testing revealed that immunization with the newly designed vaccine construct bearing the GalNAc glycocluster induced a higher titer of anti-Tn-MUC1 antibodies compared to the TACAs alone. Additionally, the antibodies obtained bind a library of tumor-associated saccharide structures on MUC1 and MUC1-positive breast cancer cells. Conjugation of a high-affinity ligand for MGL to tumor-associated MUC1 glycopeptide antigens has a synergistic impact on antibody production.
Mucin-1 (MUC1) is a highly attractive antigenic target for anticancer vaccines. Naturally existing MUC1 can contain multiple types of O-linked glycans, including the Thomsen–Friedenreich (Tf) antigen ...and the Sialyl Thomsen-nouveau (STn) antigen. In order to target these antigens as potential anticancer vaccines, MUC1 glycopeptides SAPDT*RPAP (T* is the glycosylation site) bearing the Tf and the STn antigen, respectively, have been synthesized. The bacteriophage Qβ carrier is a powerful carrier for antigen delivery. The conjugates of MUC1-Tf and -STn glycopeptides with Qβ were utilized to immunize immune-tolerant human MUC1 transgenic (MUC1.Tg) mice, which elicited superior levels of anti-MUC1 IgG antibodies with titers reaching over 2 million units. The IgG antibodies recognized a wide range of MUC1 glycopeptides bearing diverse glycans. Antibodies induced by Qβ-MUC1-Tf showed strongest binding, with MUC1-expressing melanoma B16-MUC1 cells, and effectively killed these cells in vitro. Vaccination with Qβ-MUC1-Tf first followed by tumor challenge in a lung metastasis model showed significant reductions of the number of tumor foci in the lungs of immunized mice as compared to those in control mice. This was the first time that a MUC1-Tf-based vaccine has shown in vivo efficacy in a tumor model. As such, Qβ-MUC1 glycopeptide conjugates have great potential as anticancer vaccines.
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