ABSTRACT
DC-SIGN (dendritic cell-specific ICAM-3 grabbing non-integrin) is a C-type lectin receptor (CLR) present, mainly in dendritic cells (DCs), as one of the major pattern recognition receptors (PRRs). This receptor has a relevant role in viral infection processes. Recent approaches aiming to block DC-SIGN have been presented as attractive anti-HIV strategies. DC-SIGN binds mannose or fucose-containing carbohydrates from viral proteins such as the HIV envelope glycoprotein gp120. We have previously demonstrated that multivalent dendrons bearing multiple copies of glycomimetic ligands were able to inhibit DC-SIGN-dependent HIV infection in cervical explant models. Optimization of glycomimetic ligands requires detailed characterization and analysis of their binding modes because they notably influence binding affinities. In a previous study we characterized the binding mode of DC-SIGN with ligand 1, which shows a single binding mode as demonstrated by NMR and X-ray crystallography. In this work we report the binding studies of DC-SIGN with pseudotrisaccharide 2, which has a larger affinity. Their binding was analysed by TR-NOESY and STD NMR experiments, combined with the CORCEMA-ST protocol and molecular modelling. These studies demonstrate that in solution the complex cannot be explained by a single binding mode. We describe the ensemble of ligand bound modes that best fit the experimental data and explain the higher inhibition values found for ligand 2.
Subject(s)
Cell Adhesion Molecules/chemistry , Lectins, C-Type/chemistry , Receptors, Cell Surface/chemistry , Trisaccharides/pharmacology , Binding Sites/drug effects , Cell Adhesion Molecules/metabolism , Crystallography, X-Ray , Dendritic Cells , Humans , Lectins, C-Type/metabolism , Ligands , Models, Molecular , Molecular Structure , Nuclear Magnetic Resonance, Biomolecular , Protein Binding/drug effects , Receptors, Cell Surface/metabolism , Structure-Activity Relationship , Trisaccharides/chemical synthesis , Trisaccharides/chemistryABSTRACT
Here we propose the optimization of a rapid and reproducible protocol for intracellular metabolite extraction from yeast cells and their metabolic profiling by (1)H-NMR spectroscopy. The protocol reliability has been validated through comparison between the metabolome of cells in different phases of growth or with different genetic backgrounds.
Subject(s)
Metabolomics/methods , Proton Magnetic Resonance Spectroscopy/methods , Saccharomycetales/metabolism , Metabolome , Saccharomycetales/cytology , Saccharomycetales/growth & developmentABSTRACT
The aim of this research was to evaluate the Ergovision Screener (ES) accuracy e validity by a confrontation with the conventional ophthalmic check (OC), for the medical evaluation of job fitness. A population of 100 VDU operators was considered. Each subject underwent randomly both the ES and the ophthalmic check visit. Several test carried out by the Ergovision Screener were not consistent with the conventional ophthalmic check. In a number of cases, high false positive ratio have been found, which could lead to unnecessary further examinations. For all these reasons we believe that the ES is not an appropriate instrument for the medical evaluation of job fitness.
