Ana Ardá

Total synthesis of (-)-dysithiazolamide.

The tetrachlorinated natural product (-)-dysithiazolamide was synthesized from l-glutamic acid in a convergent way, confirming the previously proposed (2S,4S,6S,8S) absolute stereochemistry.

Thermodynamic Switch in Binding of Adhesion/Growth Regulatory Human Galectin-3 to Tumor-Associated TF Antigen (CD176) and MUC1 Glycopeptides

A shift to short-chain glycans is an observed change in mucin-type O-glycosylation in premalignant and malignant epithelia. Given the evidence that human galectin-3 can interact with mucins and also weakly with free tumor-associated Thomsen-Friedenreich (TF) antigen (CD176), the study of its interaction with MUC1 (glyco)peptides is of biomedical relevance. Glycosylated MUC1 fragments that carry the TF antigen attached through either Thr or Ser side chains were synthesized using standard Fmoc-based automated solid-phase peptide chemistry. The dissociation constants (Kd) for interaction of galectin-3 and the glycosylated MUC1 fragments measured by isothermal titration calorimetry decreased up to 10 times in comparison to that of the free TF disaccharide. No binding was observed for the nonglycosylated control version of the MUC1 peptide. The most notable feature of the binding of MUC1 glycopeptides to galectin-3 was a shift from a favorable enthalpy to an entropy-driven binding process. The comparatively diminished enthalpy contribution to the free energy (ΔG) was compensated by a considerable gain in the entropic term. 1H–15N heteronuclear single-quantum coherence spectroscopy nuclear magnetic resonance data reveal contact at the canonical site mainly by the glycan moiety of the MUC1 glycopeptide. Ligand-dependent differences in binding affinities were also confirmed by a novel assay for screening of low-affinity glycan–lectin interactions based on AlphaScreen technology. Another key finding is that the glycosylated MUC1 peptides exhibited activity in a concentration-dependent manner in cell-based assays revealing selectivity among human galectins. Thus, the presentation of this tumor-associated carbohydrate ligand by the natural peptide scaffold enhances its affinity, highlighting the significance of model studies of human lectins with synthetic glycopeptides.

Low-Temperature NMR J-Based Configurational Analysis of Flexible Acyclic Systems

A new strategy for the J-based configurational analysis in acyclic systems is applied where the existence of a multiple conformer equilibrium precludes the application of the Murata methodology.

Glycans in Infectious Diseases. A Molecular Recognition Perspective

BACKGROUND From the simplest bacteria to the highest complex mammals, including humans, every single cell is covered by a dense coat of glycans. Glycans are involved in almost every biological process that takes place in our body, playing a central role in the communication between cells and their environment. Glycans are also involved in infectious diseases, which arise from the specific interaction between glycans of the pathogen cell coat and specific receptors on the host cell or vice versa. OBJECTIVE The understanding of the mechanisms governing these specific carbohydrateprotein interactions, at atomic and molecular levels, is crucial to develop new drugs able to block the infection and to avoid the disease. METHODS Recent advances in biophysical techniques allow for a complete picture of the hostpathogen infection event, unveiling the key aspects of the molecular interaction and, thus, providing an opportunity to interfere with it. CONCLUSION In this general review, we discuss some recent contributions, providing a summary of what we consider the most innovative and inspiring research lines to the field.

Fluoroacetamide Moieties as NMR Probes for molecular recognition of GlcNAc‐containing sugars: Modulation of the CH‐π Stacking Interactions by Different Fluorination Patterns

Abstract We herein propose the use of fluoroacetamide and difluoroacetamide moieties as sensitive tags for the detection of sugar–protein interactions by simple 1H and/or 19F NMR spectroscopy methods. In this process, we have chosen the binding of N,N′‐diacetyl chitobiose, a ubiquitous disaccharide fragment in glycoproteins, by wheat‐germ agglutinin (WGA), a model lectin. By using saturation‐transfer difference (STD)‐NMR spectroscopy, we experimentally demonstrate that, under solution conditions, the molecule that contained the CHF2CONH‐ moiety is the stronger aromatic binder, followed by the analogue with the CH2FCONH‐ group and the natural molecule (with the CH3CONH‐ fragment). In contrast, the molecule with the CF3CONH‐ isoster displayed the weakest intermolecular interaction (one order of magnitude weaker). Because sugar–aromatic CH–π interactions are at the origin of these observations, these results further contribute to the characterization and exploration of these forces and offer an opportunity to use them to unravel complex recognition processes.

CHAPTER 1:New Applications of High‐Resolution NMR in Drug Discovery and Development

NMR spectroscopy is among the most powerful biophysical techniques in drug discovery, its prominent position owing to the existence of a wide variety of methods taking advantage of the properties of NMR active nuclei to extract structural and dynamic information useful at all stages of the drug‐discovery process. This chapter gathers the most important state‐of‐the‐art NMR procedures used in drug discovery in both industry and academia, from screening techniques (STD, WaterLOGSY, relaxation filtering, TINS) to experiments providing structural information on both ligand and receptor in the bound state (TR‐NOESY, paramagnetic spin labeling, INPHARMA, diffusion‐based methods, SAR by NMR). Practical aspects of the application of NMR to fragment‐based drug discovery are included. Advances in the field that are currently expanding the applicability of NMR methodologies to larger and membrane‐bound systems are also discussed.

Environmental Effects Determine the Structure of Potential β-Amino Acid Based Foldamers

This work shows that hybrid peptides formed by alternating trans-2-aminocyclopentanecarboxylic acid (trans-ACPC) and trans-2-aminocyclohexanecarboxylic acid (trans-ACHC) do not fold in the solvents typically used in the study of their homo-oligomers. Only when the peptides are assayed in SDS micelles are the predicted helical structures obtained. This indicates that the environment could play an equally important role (as the backbone stereochemistry) in determining their fold, possibly by providing a sequestered environment.

Recent advances in the application of NMR methods to uncover the conformation and recognition features of glycans

This chapter is dedicated to the presentation of different examples of the application of solution NMR to the study of conformation, dynamics of sugar molecules (oligo and polysaccharides, glycopeptides and glycomimetics) and to the investigation of glycan-related molecular recognition events. Selected examples since 2012 are presented depending on the chemical nature of the sugar molecule, on the environment (free or bound) and on the nature of the receptor.

Chapter 1:Carbohydrate–Protein Interactions: A 3D View by NMR

NMR spectroscopy is a key tool for carbohydrate research. In studies with complex oligosaccharides there are limits to the amount of relevant structural information provided by these observables due to problems of signal overlapping, strong coupling and/or the scarcity of the key NOE information. Thus, there is an increasing need for additional parameters with structural information, such as residual dipolar couplings (RDCs), paramagnetic relaxation enhancements (PREs) or pseudo contact shifts (PCSs). Carbohydrates are rather flexible molecules. Therefore, NMR observables do not always correlate with a single conformer but with an ensemble of low free-energy conformers that can be accessed by thermal fluctuations. Depending on the system under study, different NMR approaches can be followed to characterize protein–carbohydrate interactions: the standard methodologies can usually be classified as “ligand-based” or “receptor-based”. The selection of the proper methodology is usually determined by the size of the receptor, the dissociation constant of the complex (KD), the availability of the labelled protein (15N, 13C) and the access to soluble receptors at enough concentration for NMR measurements.