Ana Luísa M. de Carvalho

Interactions of glycan-specific epitopes to human lectin receptors represent novel immune checkpoints for investigating cancer and infection diseases. By employing a multidisciplinary approach that combines isothermal titration calorimetry, NMR spectroscopy, molecular dynamics simulations, and X-ray crystallography, we disclosed the molecular determinants that govern the recognition of the tumour and pathogenic glycobiomarker LacdiNAc (GalNAc?1-4GlcNAc, LDN), including their comparison with the ubiquitous LacNAc epitope (Gal?1-4GlcNAc, LN), by two human immune-related lectins, galectin-3 (hGal-3) and the macrophage galactose C-type lectin (hMGL). A different mechanism of binding and interactions is observed for the hGal-3/LDN and hMGL/LDN complexes, which explains the remarkable difference in the binding specificity of LDN and LN by these two lectins. The new structural clues reported herein are fundamental for the chemical design of mimetics targeting hGal-3/hMGL recognition process.

Overexpression of the Thomsen-Friedenreich (TF) antigen in cell membrane proteins occurs in 90% of adenocarcinomas. Additionally, the binding of the TF-antigen to human galectin-3 (Gal-3), also frequently overexpressed in malignancy, promotes cancer progression and metastasis. In this context, structures that interfere with this specific interaction display the potential to prevent cancer metastasis. Herein, a multidisciplinary approach, combining the optimized synthesis of a TF-antigen mimetic with NMR, X-ray crystallography methods and isothermal titration calorimetry assays has been employed to unravel the molecular structural details that govern the Gal-3/TF-mimetic interaction. The TF-mimetic presents a binding affinity for Gal-3 similar to the TF-natural antigen and retains the binding epitope and the bioactive conformation observed for the native antigen. Furthermore, from a thermodynamic perspective a decrease in the enthalpic contribution was observed for the Gal-3/TF-mimetic complex, however this behaviour is compensated by a favourable entropy gain. From a structural perspective, these results establish our TF-mimetic as a scaffold to design multivalent solutions to potentially interfere with Gal-3 aberrant interactions and likely be used to hamper Gal-3-mediated cancer cells adhesion and metastasis.

The functional and biological significance of the selected CASP12 targets are described by the authors of the structures. The crystallographers discuss the most interesting structural features of the target proteins and assess whether these features were correctly reproduced in the predictions submitted to the CASP12 experiment. This article is protected by copyright. All rights reserved.

We show that a physical trigger, a non-ionizing infrared (IR) radiation at wavelengths strongly absorbed by liquid water, can be used to induce and kinetically control protein (periodic) self-assembly in solution. This phenomenon is explained by considering the effect of IR light on the structuring of protein interfacial water. Our results indicate that the IR radiation can promote enhanced mutual correlations of water molecules in the protein hydration shell. We report on the radiation-induced increase in both the strength and cooperativeness of H-bonds. The presence of a structured dipolar hydration layer can lead to attractive interactions between like-charged biomacromolecules in solution (and crystal nucleation events). Furthermore, our study suggests that enveloping the protein within a layer of structured solvent (an effect enhanced by IR light) can prevent the protein non-specific aggregation favoring periodic self-assembly. Recognizing the ability to affect protein-water interactions by means of IR radiation may have important implications for biological and bio-inspired systems.

Nucleation is a critical step determining the outcome of the entire crystallization process. Finding an effective nucleant for protein crystallization is of utmost importance for structural biology. The latter relies on good-quality crystals to solve the three-dimensional structures of macromolecules. In this study we show that crystalline barium sulfate (BaSO4) with an etched and/or ionic liquid (IL)-functionalized surface (1) can induce protein nucleation at concentrations well below the concentration needed to promote crystal growth under control conditions, (2) can shorten the nucleation time, (3) can increase the growth rate, and finally (4) may help to improve the protein crystal morphology. These effects were shown for lysozyme, RNase A, trypsin, proteinase K, myoglobin, and hemoglobin. Therefore, the use of BaSO4 particles enables us to reduce the amount of protein in crystallization trials and increases the chance of obtaining protein crystals of the desired quality. In the context of the underlying mechanism, it is shown that the protein-solid contact formation is governed by the interaction of the polar compartments of the biomacromolecule with the support. The tendency of a protein to concentrate near the solid surface is enhanced by both the hydrophobicity of the protein and that of the surface (tuned by the functionalizing IL). These mechanisms of interaction of biomacromolecules with inorganic hydrophilic solids correspond to the principles of amphiphilic IL-mineral interactions.