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10 p.-6 fig.-3 tb., Functional pairing between cellular glyco-con-ju-gates and tissue lectins like galectins has wide (patho)physio-logical significance. Their study is facilitated by non-hydrolyzable deri-va-tives of the natural O-glycans, such as S- and Se-glycosides. The latter en-able extensive analyses by specific 77 Se NMR spectroscopy, but still remain under-explored. By the example of seleno--digalac-to-side (SeDG) and the human galectins 1 and 3, we evaluate diverse 77 Se NMR detec-tion methods and pro-pose selec-tive 1 H, 77 Se hetero-nuclear Hartmann-Hahn transfer for efficient use in com-peti-tive NMR screening against a seleno-glycoside spy ligand. By fluores-cence aniso-tropy, circular dichroism, and isothermal titra-tion calori-metry (ITC) we show that affinity and thermodynamics of SeDG bind-ing by galectins are similar to thiodi-galac-toside (TDG) and N-acetyl-lactos-amine (LacNAc), con-firm-ing that Se substitution has no major impact. ITC data in D 2 O vs. H 2 O are similar for TDG and LacNAc binding by both galectins, but a solvent effect, indicating solvent rearrangement at the binding site, is hinted for SeDG and clearly observed for LacNAc dimers with extend-ed chain length., We are much indebted to L. Szilágyi (University of Debrecen,Hungary) for kindly providing selenoglycosides for this study. We are grateful for inspiring discussions to Drs. B. Friday, A. Leddoz and A. W. L. Nose as well as for generous funding from the Spanish Ministry of Economy and Competitiveness (grant BFU2016-77835-R; to A.R.), the Department of Industry, Tourism and Trade of the Government of the Autonomous Community of the Basque Country (Elkartek BG2019; to T.D.), the Severo Ochoa Excellence Accreditation from MCIU (SEV-2016-0644; to T.D.), the Science Foundation of Ireland (grants 13/IA/1959 and 16/RC/3889;to S.O.), National Institute of Health (NIH) grant CA242351 to M.Cudic, and the COST Action CA 18103 (InnoGly; to H.-J.G.).

Glycan-protein interactions play an important role in a broad range of physiological processes, raising interest to elucidate the structural interplay. Yet, their dynamic nature limits the analysis by crystallography, whereas NMR spectroscopy suffers from the low (1) H dispersion of glycans. Therefore, their sparse fluorination and NMR screening by 1D Saturation Transfer Difference with relay to (19) F (STDreF) was previously proposed to exploit the superior dispersion in (19) F NMR spectroscopy. A new 2D STD-TOCSYreF experiment is presented here that enables comprehensive epitope mapping of fluorinated glycans by combining the spectral resolution of (19) F with the spatial resolution and coverage of (1) H. For an illustration, the 2-deoxy-2-fluoro derivative of the N-glycan core trimannoside was synthesised and its recognition of Pisum sativum agglutinin by either of the two terminal mannose residues was confirmed. Going beyond the crystallographic information, the 2D STD-TOCSYreF spectrum moreover visualised collateral contacts from the branching mannose and allowed to assess the ratio of both co-existing binding modes through the alpha1,3- (67 %) and alpha1,6-linked (33 %) terminal mannose moieties.

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.

3 páginas, 2 figuras -- PAGS nros. 5666-5668, As a novel approach for studying carbohydrate-lectin interactions spectroscopically, we combine the resolution and specificity of (19)F-detected NMR spectroscopy with the sensitivity of the saturation transfer difference (STD) technique. The resulting background-free (19)F-STD spectra open a promising perspective for broad application with medical relevance, for example, in drug discovery, Financial support by a bizkaia: Xede grant, an EC Marie Curie Research Training network (MRTN-CT2005-019561), the Ministry of Science and Innovation of Spain (CTQ2006- 10874-C02-01)