Molecular Recognition in C-Type Lectins: The Cases of DC-SIGN, Langerin, MGL, and L-Sectin.

Carbohydrates play a pivotal role in intercellular communication processes. In particular, glycan antigens are key for sustaining homeostasis, helping leukocytes to distinguish damaged tissues and invading pathogens from healthy tissues. From a structural perspective, this cross-talk is fairly complex, and multiple membrane proteins guide these recognition processes, including lectins and Toll-like receptors. Since the beginning of this century, lectins have become potential targets for therapeutics for controlling and/or avoiding the progression of pathologies derived from an incorrect immune outcome, including infectious processes, cancer, or autoimmune diseases. Therefore, a detailed knowledge of these receptors is mandatory for the development of specific treatments. In this review, we summarize the current knowledge about four key C-type lectins whose importance has been steadily growing in recent years, focusing in particular on how glycan recognition takes place at the molecular level, but also looking at recent progresses in the quest for therapeutics.

Fluorinated Carbohydrates as Lectin Ligands: Simultaneous Screening of a Monosaccharide Library and Chemical Mapping by 19F NMR Spectroscopy.

Molecular recognition of carbohydrates is a key step in essential biological processes. Carbohydrate receptors can distinguish monosaccharides even if they only differ in a single aspect of the orientation of the hydroxyl groups or harbor subtle chemical modifications. Hydroxyl-by-fluorine substitution has proven its merits for chemically mapping the importance of hydroxyl groups in carbohydrate-receptor interactions. 19F NMR spectroscopy could thus be adapted to allow contact mapping together with screening in compound mixtures. Using a library of fluorinated glucose (Glc), mannose (Man), and galactose (Gal) derived by systematically exchanging every hydroxyl group by a fluorine atom, we developed a strategy combining chemical mapping and 19F NMR T2 filtering-based screening. By testing this strategy on the proof-of-principle level with a library of 13 fluorinated monosaccharides to a set of three carbohydrate receptors of diverse origin, i.e. the human macrophage galactose-type lectin, a plant lectin, Pisum sativum agglutinin, and the bacterial Gal-/Glc-binding protein from Escherichia coli, it became possible to simultaneously define their monosaccharide selectivity and identify the essential hydroxyls for interaction.

Unraveling Sugar Binding Modes to DC-SIGN by Employing Fluorinated Carbohydrates.

A fluorine nuclear magnetic resonance (19F-NMR)-based method is employed to assess the binding preferences and interaction details of a library of synthetic fluorinated monosaccharides towards dendritic cell-specific intercellular adhesion molecule 3-grabbing non-integrin (DC-SIGN), a lectin of biomedical interest, which is involved in different viral infections, including HIV and Ebola, and is able to recognize a variety of self- and non-self-glycans. The strategy employed allows not only screening of a mixture of compounds, but also obtaining valuable information on the specific sugar-protein interactions. The analysis of the data demonstrates that monosaccharides Fuc, Man, Glc, and Gal are able to bind DC-SIGN, although with decreasing affinity. Moreover, a new binding mode between Man moieties and DC-SIGN, which might have biological implications, is also detected for the first time. The combination of the 19F with standard proton saturation transfer difference (1H-STD-NMR) data, assisted by molecular dynamics (MD) simulations, permits us to successfully define this new binding epitope, where Man coordinates a Ca2+ ion of the lectin carbohydrate recognition domain (CRD) through the axial OH-2 and equatorial OH-3 groups, thus mimicking the Fuc/DC-SIGN binding architecture.

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 19F-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. A novel 2D-STD-TOCSYreF NMR experiment was employed to obtain the experimental STD NMR intensities, while the Complete Relaxation Matrix Analysis (CORCEMA-ST) was used to predict that expected for an ensemble of geometries extracted from extensive MD simulations. Then, an in-house built computer program was devised to find the ensemble of structures that provide the best fit between the theoretical and the observed STD data. Remarkably, the experimental STD profiles obtained for the ligand/DC-SIGN complex could not be satisfactorily explained by a single binding mode, but rather with a combination of different modes coexisting in solution. Therefore, the method provides a precise view of those ligand-receptor complexes present in solution.

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 LeX. 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 LeX. Herein, we have studied the interaction in solution between DC-SIGN and the blood group A and B antigens, to get insights into the atomic details of such interaction. With a combination of different NMR experiments, we demonstrate that the Fuc coordinates the primary Ca2+ ion with a single binding mode through 3-OH and 4-OH. The terminal αGal/αGalNAc affords marginal direct polar contacts with the protein, but provides a hydrophobic hook in which V351 of the lectin perfectly fits. Moreover, we have found that αGal, but not αGalNAc, is a weak binder itself for DC-SIGN, which could endow an additional binding mode for the blood group B antigen, but not for blood group A.