Molecular Recognition of Glycan-Bearing Glycomacromolecules Presented at Membrane Surfaces by Lectins: An NMR View.

Lectin-glycan interactions are at the heart of a multitude of biological events. Glycans are usually presented in a multivalent manner on the cell surface as part of the so-called glycocalyx, where they interact with other entities. This multivalent presentation allows us to overcome the typical low affinities found for individual glycan-lectin interactions. Indeed, the presentation of glycans may drastically impact their binding by lectins, highly affecting the corresponding binding affinity and even selectivity. In this context, we herein present the study of the interaction of a variety of homo- and heteromultivalent lactose-functionalized glycomacromolecules and their lipid conjugates with two human galectins. We have employed as ligands the glycomacromolecules, as well as liposomes decorated with those structures, to evaluate their interactions in a cell-mimicking environment. Key details of the interaction have been unravelled by NMR experiments, both from the ligand and receptor perspectives, complemented by cryo-electron microscopy methods and molecular dynamics simulations.

Assessing the Mobility of Severe Acute Respiratory Syndrome Coronavirus-2 Spike Protein Glycans by Structural and Computational Methods.

Two years after its emergence, the coronavirus disease-2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) remains difficult to control despite the availability of several vaccines. The extensively glycosylated SARS-CoV-2 spike (S) protein, which mediates host cell entry by binding to the angiotensin converting enzyme 2 (ACE2) through its receptor binding domain (RBD), is the major target of neutralizing antibodies. Like to many other viral fusion proteins, the SARS-CoV-2 spike protein utilizes a glycan shield to thwart the host immune response. To grasp the influence of chemical signatures on carbohydrate mobility and reconcile the cryo-EM density of specific glycans we combined our cryo-EM map of the S ectodomain to 4.1 Å resolution, reconstructed from a limited number of particles, and all-atom molecular dynamics simulations. Chemical modifications modeled on representative glycans (defucosylation, sialylation and addition of terminal LacNAc units) show no significant influence on either protein shielding or glycan flexibility. By estimating at selected sites the local correlation between the full density map and atomic model-based maps derived from molecular dynamics simulations, we provide insight into the geometries of the α-Man-(1→3)-[α-Man-(1→6)-]-ß-Man-(1→4)-ß-GlcNAc(1→4)-ß-GlcNAc core common to all N-glycosylation sites.

Bacteriophage PRD1 as a nanoscaffold for drug loading.

Viruses are very attractive biomaterials owing to their capability as nanocarriers of genetic material. Efforts have been made to functionalize self-assembling viral protein capsids on their exterior or interior to selectively take up different payloads. PRD1 is a double-stranded DNA bacteriophage comprising an icosahedral protein outer capsid and an inner lipidic vesicle. Here, we report the three-dimensional structure of PRD1 in complex with the antipsychotic drug chlorpromazine (CPZ) by cryo-electron microscopy. We show that the jellyrolls of the viral major capsid protein P3, protruding outwards from the capsid shell, serve as scaffolds for loading heterocyclic CPZ molecules. Additional X-ray studies and molecular dynamics simulations show the binding modes and organization of CPZ molecules when complexed with P3 only and onto the virion surface. Collectively, we provide a proof of concept for the possible use of the lattice-like organisation and the quasi-symmetric morphology of virus capsomers for loading heterocyclic drugs with defined properties.

Cross-Linking Effects Dictate the Preference of Galectins to Bind LacNAc-Decorated HPMA Copolymers.

The interaction of multi-LacNAc (Galß1-4GlcNAc)-containing N-(2-hydroxypropyl) methacrylamide (HPMA) copolymers with human galectin-1 (Gal-1) and the carbohydrate recognition domain (CRD) of human galectin-3 (Gal-3) was analyzed using NMR methods in addition to cryo-electron-microscopy and dynamic light scattering (DLS) experiments. The interaction with individual LacNAc-containing components of the polymer was studied for comparison purposes. For Gal-3 CRD, the NMR data suggest a canonical interaction of the individual small-molecule bi- and trivalent ligands with the lectin binding site and better affinity for the trivalent arrangement due to statistical effects. For the glycopolymers, the interaction was stronger, although no evidence for forming a large supramolecule was obtained. In contrast, for Gal-1, the results indicate the formation of large cross-linked supramolecules in the presence of multivalent LacNAc entities for both the individual building blocks and the polymers. Interestingly, the bivalent and trivalent presentation of LacNAc in the polymer did not produce such an increase, indicating that the multivalency provided by the polymer is sufficient for triggering an efficient binding between the glycopolymer and Gal-1. This hypothesis was further demonstrated by electron microscopy and DLS methods.

Superimposition of Viral Protein Structures: A Means to Decipher the Phylogenies of Viruses.

Superimposition of protein structures is key in unravelling structural homology across proteins whose sequence similarity is lost. Structural comparison provides insights into protein function and evolution. Here, we review some of the original findings and thoughts that have led to the current established structure-based phylogeny of viruses: starting from the original observation that the major capsid proteins of plant and animal viruses possess similar folds, to the idea that each virus has an innate "self". This latter idea fueled the conceptualization of the PRD1-adenovirus lineage whose members possess a major capsid protein (innate "self") with a double jelly roll fold. Based on this approach, long-range viral evolutionary relationships can be detected allowing the virosphere to be classified in four structure-based lineages. However, this process is not without its challenges or limitations. As an example of these hurdles, we finally touch on the difficulty of establishing structural "self" traits for enveloped viruses showcasing the coronaviruses but also the power of structure-based analysis in the understanding of emerging viruses.