Crystal structures of peanut lectin in the presence of synthetic β-N- and β-S-galactosides disclose evidences for the recognition of different glycomimetic ligands

Abstract

ORCID:https://orcid.org/0000-0002-5448-5483. # equally contributed to this work Keywords Carbohydrate−protein interactions, peanut agglutinin, glycoclusters, multivalent ligands, lectin structure, X-ray crystallography, isothermal titration calorimetry, molecular dynamics. AbstractCarbohydrate−lectin interactions are involved in important cellular recognition processes, including viral and bacterial infections, inflammation, and tumor metastasis. Hence, the structural studies of lectin-synthetic glycan complexes are essential for understanding the lectin recognition processes and the further design of promising chemotherapeutics that interfere with sugar-lectin interactions.Plant lectins are excellent models for the study of the molecular recognition process. Among them, peanut lectin (PNA) is highly relevant in the glycobiology field, because of its specificity for β-galactosides, showing high affinity towards the Thomsen-Friedenreich (TF) antigen, a well-known tumor-associated carbohydrate antigen. Given this specificity, PNA is one of the most frequently used molecular probes for the recognition of tumor cell-surface O-glycans. Thus, it has been extensively used in glycobiology for inhibition studies with a variety of βgalactoside and β-lactoside ligands. Herein, crystal structures of PNA are reported in complex with six novel synthetic hydrolytically stable β-N-and β-S-galactosides. These complexes, along with computational simulations, disclosed key molecular binding interactions of the different sugars to PNA at the atomic level, revealing the role of specific water molecules in the protein-ligand recognition. Furthermore, binding affinity studies measured by isothermal titration calorimetry showed dissociation constant values in the micromolar range, as well as a positive glycoside cluster effect in terms of affinity in the case of the divalent compounds. Taken together, this work provides qualitative structural rationale for the upcoming synthesis of optimized glycoclusters, designed for the study of lectinmediated biological processes. The understanding of the recognition of β-N-and β-Sgalactosides with PNA represents a benchmark in protein-carbohydrate interactions since they are novel synthetic ligands not belonging to the family of O-linked glycosides.