Evolution of human H3N2 influenza virus receptor specificity has substantially expanded the receptor-binding domain site.

Hemagglutinins (HAs) from human influenza viruses descend from avian progenitors that bind α2-3-linked sialosides and must adapt to glycans with α2-6-linked sialic acids on human airway cells to transmit within the human population. Since their introduction during the 1968 pandemic, H3N2 viruses have evolved over the past five decades to preferentially recognize human α2-6-sialoside receptors that are elongated through addition of poly-LacNAc. We show that more recent H3N2 viruses now make increasingly complex interactions with elongated receptors while continuously selecting for strains maintaining this phenotype.

Revealing the Specificity of Human H1 Influenza A Viruses to Complex -Glycans.

Influenza virus infection remains a threat to human health since viral hemagglutinins are constantly drifting, escaping infection and vaccine-induced antibody responses. Viral hemagglutinins from different viruses display variability in glycan recognition. In this context, recent H3N2 viruses have specificity for α2,6 sialylated branched -glycans with at least three -acetyllactosamine units (tri-LacNAc).

Synthesis and Characterization of a Novel Resveratrol Xylobioside Obtained Using a Mutagenic Variant of a GH10 Endoxylanase.

Resveratrol is a natural polyphenol with antioxidant activity and numerous health benefits. However, in vivo application of this compound is still a challenge due to its poor aqueous solubility and rapid metabolism, which leads to an extremely low bioavailability in the target tissues. In this work, rXynSOS-E236G glycosynthase, designed from a GH10 endoxylanase of the fungus was used to glycosylate resveratrol by using xylobiosyl-fluoride as a sugar donor.

A Fungal Versatile GH10 Endoxylanase and Its Glycosynthase Variant: Synthesis of Xylooligosaccharides and Glycosides of Bioactive Phenolic Compounds.

The study of endoxylanases as catalysts to valorize hemicellulosic residues and to obtain glycosides with improved properties is a topic of great industrial interest. In this work, a GH10 β-1,4-endoxylanase (XynSOS), from the ascomycetous fungus , has been heterologously produced in , purified, and characterized. rXynSOS is a highly glycosylated monomeric enzyme of 53 kDa that contains a functional CBM1 domain and shows its optimal activity on azurine cross-linked (AZCL)-beechwood xylan at 70 °C and pH 5.

Conformational and Structural Characterization of Carbohydrates and their Interactions Studied by NMR.

Carbohydrates, either free or as glycans conjugated with other biomolecules, participate in a plethora of essential biological processes. Their apparent simplicity in terms of chemical functionality hides an extraordinary diversity and structural complexity. Deeply deciphering at the atomic level their structures is essential to understand their biological function and activities, but it is still a challenging task in need of complementary approaches and no generalized procedures are available to address the study of such complex, natural glycans.

Structural basis for recognition of bacterial cell wall teichoic acid by pseudo-symmetric SH3b-like repeats of a viral peptidoglycan hydrolase.

Endolysins are bacteriophage-encoded peptidoglycan hydrolases targeting the cell wall of host bacteria their cell wall-binding domains (CBDs). The molecular basis for selective recognition of surface carbohydrate ligands by CBDs remains elusive. Here, we describe, in atomic detail, the interaction between the phage endolysin domain CBD500 and its cell wall teichoic acid (WTA) ligands.

Crystal Structure of the Carbohydrate Recognition Domain of the Human Macrophage Galactose C-Type Lectin Bound to GalNAc and the Tumor-Associated Tn Antigen.

The human macrophage galactose lectin (MGL) is an endocytic type II transmembrane receptor expressed on immature monocyte-derived dendritic cells and activated macrophages and plays a role in modulating the immune system in response to infections and cancer. MGL contains an extracellular calcium-dependent (C-type) carbohydrate recognition domain (CRD) that specifically binds terminal acetylgalactosamine glycan residues such as the Tn and sialyl-Tn antigens found on tumor cells, as well as other and glycans displayed on certain viruses and parasites. Even though the glycan specificity of MGL is known and several binding glycoproteins have been identified, the molecular basis for substrate recognition has remained elusive due to the lack of high-resolution structures.

Synthesis and Evaluation of Novel Iminosugars Prepared from Natural Amino Acids.

Cyclopropanated iminosugars have a locked conformation that may enhance the inhibitory activity and selectivity against different glycosidases. We show the synthesis of new cyclopropane-containing piperidines bearing five stereogenic centers from natural amino acids l-serine and l-alanine. Those prepared from the latter amino acid may mimic l-fucose, a natural-occurring monosaccharide involved in many molecular recognition events.

Thioglycoligase derived from fungal GH3 β-xylosidase is a multi-glycoligase with broad acceptor tolerance.

The synthesis of customized glycoconjugates constitutes a major goal for biocatalysis. To this end, engineered glycosidases have received great attention and, among them, thioglycoligases have proved useful to connect carbohydrates to non-sugar acceptors. However, hitherto the scope of these biocatalysts was considered limited to strong nucleophilic acceptors.

The Interaction of Fluorinated Glycomimetics with DC-SIGN: Multiple Binding Modes Disentangled by the Combination of NMR Methods and MD Simulations.

Fluorinated glycomimetics are frequently employed to study and eventually modulate protein-glycan interactions. However, complex glycans and their glycomimetics may display multiple binding epitopes that enormously complicate the access to a complete picture of the protein-ligand complexes. We herein present a new methodology based on the synergic combination of experimental F-based saturation transfer difference (STD) NMR data with computational protocols, applied to analyze the interaction between DC-SIGN, a key lectin involved in inflammation and infection events with the trifluorinated glycomimetic of the trimannoside core, ubiquitous in human glycoproteins.

Dissecting the Essential Role of Anomeric β-Triflates in Glycosylation Reactions.

Glycosylations promoted by triflate-generating reagents are widespread synthetic methods for the construction of glycosidic scaffolds and glycoconjugates of biological and chemical interest. These processes are thought to proceed with the participation of a plethora of activated high energy intermediates such as the α- and β-glycosyl triflates, or even increasingly unstable glycosyl oxocarbenium-like species, among which only α-glycosyl triflates have been well characterized under representative reaction conditions. Interestingly, the remaining less accessible intermediates, yet to be experimentally described, seem to be particularly relevant in α-selective processes, involving weak acceptors.

A glucotolerant β-glucosidase from the fungus Talaromyces amestolkiae and its conversion into a glycosynthase for glycosylation of phenolic compounds.

Background: The interest for finding novel β-glucosidases that can improve the yields to produce second-generation (2G) biofuels is still very high. One of the most desired features for these enzymes is glucose tolerance, which enables their optimal activity under high-glucose concentrations. Besides, there is an additional focus of attention on finding novel enzymatic alternatives for glycoside synthesis, for which a mutated version of glycosidases, named glycosynthases, has gained much interest in recent years.

Exploiting xylan as sugar donor for the synthesis of an antiproliferative xyloside using an enzyme cascade.

Background: Currently, industrial societies are seeking for green alternatives to conventional chemical synthesis. This demand has merged with the efforts to convert lignocellulosic biomass into value-added products. In this context, xylan, as one of main components of lignocellulose, has emerged as a raw material with high potential for advancing towards a sustainable economy.

A top-down chemo-enzymatic approach towards N-acetylglucosamine-N-acetylmuramic oligosaccharides: Chitosan as a reliable template.

An unprecedented approach towards oligosaccharides containing N-acetylglucosamine-N-acetylmuramic (NAG-NAM) units was developed. These novel bacterial cell wall surrogates were obtained from chitosan via a top down approach involving both chemical and enzymatic reactions. The chemical modification of chitosan using a molecular clamp based strategy, allowed obtaining N-acetylglucosamine-N-acetylmuramic (NAG-NAM) containing oligomers.

Molecular Insights into DC-SIGN Binding to Self-Antigens: The Interaction with the Blood Group A/B Antigens.

The dendritic cell-specific intracellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN) is an important receptor of the immune system. Besides its role as pathogen recognition receptor (PRR), it also interacts with endogenous glycoproteins through the specific recognition of self-glycan epitopes, like Le. However, this lectin represents a paradigmatic case of glycan binding promiscuity, and it also has been shown to recognize antigens with α1-α2 linked fucose, such as the histo blood group antigens, with similar affinities to Le.

Transglycosylation products generated by Talaromyces amestolkiae GH3 β-glucosidases: effect of hydroxytyrosol, vanillin and its glucosides on breast cancer cells.

Background: Transglycosylation represents one of the most promising approaches for obtaining novel glycosides, and plant phenols and polyphenols are emerging as one of the best targets for creating new molecules with enhanced capacities. These compounds can be found in diet and exhibit a wide range of bioactivities, such as antioxidant, antihypertensive, antitumor, neuroprotective and anti-inflammatory, and the eco-friendly synthesis of glycosides from these molecules can be a suitable alternative for increasing their health benefits.

Results: Transglycosylation experiments were carried out using different GH3 β-glucosidases from the fungus Talaromyces amestolkiae.

Minimizing the Entropy Penalty for Ligand Binding: Lessons from the Molecular Recognition of the Histo Blood-Group Antigens by Human Galectin-3.

Ligand conformational entropy plays an important role in carbohydrate recognition events. Glycans are characterized by intrinsic flexibility around the glycosidic linkages, thus in most cases, loss of conformational entropy of the sugar upon complex formation strongly affects the entropy of the binding process. By employing a multidisciplinary approach combining structural, conformational, binding energy, and kinetic information, we investigated the role of conformational entropy in the recognition of the histo blood-group antigens A and B by human galectin-3, a lectin of biomedical interest.

Increase of Redox Potential during the Evolution of Enzymes Degrading Recalcitrant Lignin.

To investigate how ligninolytic peroxidases acquired the uniquely high redox potential they show today, their ancestors were resurrected and characterized. Unfortunately, the transient Compounds I (CI) and II (CII) from peroxide activation of the enzyme resting state (RS) are unstable. Therefore, the reduction potentials (E°') of the three redox couples (CI/RS, CI/CII and CII/RS) were estimated (for the first time in a ligninolytic peroxidase) from equilibrium concentrations analyzed by stopped-flow UV/Vis spectroscopy.

Enzymatic fine-tuning for 2-(6-hydroxynaphthyl) β-D-xylopyranoside synthesis catalyzed by the recombinant β-xylosidase BxTW1 from Talaromyces amestolkiae.

Background: Glycosides are compounds displaying crucial biological roles and plenty of applications. Traditionally, these molecules have been chemically obtained, but its efficient production is limited by the lack of regio- and stereo-selectivity of the chemical synthesis. As an interesting alternative, glycosidases are able to catalyze the formation of glycosides in a process considered green and highly selective.

Chemometric Analysis of Bacterial Peptidoglycan Reveals Atypical Modifications That Empower the Cell Wall against Predatory Enzymes and Fly Innate Immunity.

Peptidoglycan is a fundamental structure for most bacteria. It contributes to the cell morphology and provides cell wall integrity against environmental insults. While several studies have reported a significant degree of variability in the chemical composition and organization of peptidoglycan in the domain Bacteria, the real diversity of this polymer is far from fully explored.

Finding the Right Candidate for the Right Position: A Fast NMR-Assisted Combinatorial Method for Optimizing Nucleic Acids Binders.

Development of strong and selective binders from promiscuous lead compounds represents one of the most expensive and time-consuming tasks in drug discovery. We herein present a novel fragment-based combinatorial strategy for the optimization of multivalent polyamine scaffolds as DNA/RNA ligands. Our protocol provides a quick access to a large variety of regioisomer libraries that can be tested for selective recognition by combining microdialysis assays with simple isotope labeling and NMR experiments.

The Y9P Variant of the Titin I27 Module: Structural Determinants of Its Revisited Nanomechanics.

The titin I27 module from human cardiac titin has become a standard in protein nanomechanics. A proline-scanning study of its mechanical clamp found three mechanically hypomorphic mutants and a paradoxically hypermorphic mutant (I27Y9P). Both types of mutants have been commonly used as substrates of several protein unfoldase machineries in studies relating protein mechanostability to translocation or degradation rates.