Langerin-Heparin Interaction: Two Binding Sites for Small and Large Ligands as revealed by a combination of NMR Spectroscopy and Cross-Linking Mapping Experiments

Muñoz-García, Juan C., Chabrol, Eric, Vives, Romain R., Thomas, Aline, de Paz, José L, Rojo, Javier, Imberty, Anne, Fieschi, Franck, Nieto, Pedro M. and Angulo, Jesus (2015) Langerin-Heparin Interaction: Two Binding Sites for Small and Large Ligands as revealed by a combination of NMR Spectroscopy and Cross-Linking Mapping Experiments. Journal of the American Chemical Society, 137 (12). pp. 4100-4110. ISSN 0002-7863

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Abstract

Langerin is a C-type lectin present on Langerhans cells that mediates capture of pathogens in a carbohydrate-dependent manner, leading to subsequent internalization and elimination in the cellular organelles called Birbeck granules. This mechanism mediated by langerin was shown to constitute a natural barrier for HIV-1 particle transmission. Besides interacting specifically with high mannose and fucosylated neutral carbohydrate structures, langerin has the ability to bind sulfated carbohydrate ligands as 6-sulfated galactosides in the Ca2+ dependent binding site. Very recently langerin was demonstrated to interact with sulfated glycosaminoglycans (GAGs), in a Ca2+ independent way, resulting in the proposal of a new binding site for GAGs. Based on those results, we have conducted a structural study of the interactions of small heparin (HEP) like oligosaccharides with langerin in solution. Heparin-bead cross-linking experiments, an approach specifically designed to identify HEP/HS binding sites in proteins were first carried out and experimentally validated the previously proposed model for the interaction of Lg ECD with 6 kDa HEP. High-resolution NMR studies of a set of 8 synthetic HEP-like trisaccharides harboring different sulfation patterns demonstrated that all of them bound to langerin in a Ca2+ dependent way. The binding epitopes were determined by STD NMR and the bound conformations by transferred NOESY experiments. These experimental data were combined with docking and molecular dynamics and resulted in the proposal of a binding mode characterized by the coordination of calcium by the two equatorial hydroxyl groups OH3 and OH4 at the non-reducing end. The binding also includes the carboxylate group at the adjacent iduronate residue. Such epitope is shared by all the 8 ligands, explaining the absence of any impact on binding from their differences in substitution pattern. Finally, in contrast to the small trisaccharides, we demonstrated that a longer HEP-like hexasaccharide, bearing an additional O-sulfate group at the non-reducing end, which precludes binding to the Ca2+ site, interacts with langerin in the previously identified Ca2+ independent binding site.

Item Type: Article
Faculty \ School: Faculty of Science > School of Pharmacy
Depositing User: Pure Connector
Date Deposited: 08 Apr 2015 10:34
Last Modified: 22 Jul 2020 00:12
URI: https://ueaeprints.uea.ac.uk/id/eprint/53092
DOI: 10.1021/ja511529x

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