Exploring Glycan-Lectin Interactions in Natural-Like Environments: A View Using NMR Experiments Inside Cell and on Cell Surface.

Glycan-mediated molecular recognition events are essential for life. NMR is widely used to monitor glycan binding to lectins in solution using isolated glycans and lectins. In this context, we herein explore diverse NMR methodologies, from both the receptor and ligand perspectives, to monitor glycan-lectin interactions under experimental conditions mimicking the native milieu inside cells and on cell surface.

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.

Synthetic Zwitterionic Streptococcus pneumoniae Type 1 Oligosaccharides Carrying Labile O-Acetyl Esters.

We herein report the first total synthesis of the Streptococcus pneumoniae serotype 1 (Sp1) oligosaccharide, a unique zwitterionic capsular polysaccharide carrying labile O-acetyl esters. The target oligosaccharides, featuring rare α-2,4-diamino-2,4,6-trideoxy galactose (AAT) and α-galacturonic acids, were assembled up to the 9-mer level, in a highly stereoselective manner using trisaccharide building blocks. The lability of the O-acetyl esters imposed a careful deprotection scheme to prevent migration and hydrolysis.

NMR Investigation of Protein-Carbohydrate Interactions: The Recognition of Glycans by Galectins Engineered with Fluorotryptophan Residues.

Fluorine ( F) incorporation into glycan-binding proteins (lectins) has been achieved and exploited to monitor the binding to carbohydrate ligands by nuclear magnetic resonance (NMR) spectroscopy. Galectins are a family of lectins that bind carbohydrates, generally with weak affinities, through a combination of intermolecular interactions including a key CH-π stacking involving a conserved tryptophan residue. Herein, Galectin-3 (Gal3) and Galectin-8 (Gal8) with one and two carbohydrate recognition domains (CRDs), respectively, were selected.

Oligosaccharide Presentation Modulates the Molecular Recognition of Glycolipids by Galectins on Membrane Surfaces.

Galectins are a family of glycan binding proteins that stand out for the wide range of biological phenomena in which they are involved. Most galectin functions are associated with their glycan binding capacities, which are generally well characterized at the oligosaccharide level, but not at the glycoprotein or glycolipid level. Glycolipids form the part of cell membranes where they can act as galectin cellular receptors.

Microwave-assisted palladium catalysed C-H acylation with aldehydes: synthesis and diversification of 3-acylthiophenes.

The use of MW allows the efficient palladium(II)-catalysed C-3 acylation of thiophenes with aldehydes C(sp)-H activation for the synthesis of (cyclo)alkyl/aryl thienyl ketones (43 examples). Compared to standard thermal conditions, the use of MW reduces the reaction time (15 to 30 min . 1 to 3 hours), leading to improved yields of the ketones (up to 92%).

Pd(II)-Catalyzed Fujiwara-Moritani Reactions for the Synthesis and Functionalization of Substituted Coumarins.

Highly substituted coumarins, privileged and versatile scaffolds for bioactive natural products and fluorescence imaging, are obtained via a Pd(II)-catalyzed direct C-H alkenylation reaction (Fujiwara-Moritani reaction), which has emerged as a powerful tool for the construction and functionalization of heterocyclic compounds because of its chemical versatility and its environmental advantages. Thus, a selective 6- cyclization led to 4-substituted coumarins in moderate yields. Selected examples have been further functionalized in C3 through a second intermolecular C-H alkenylation reaction to give coumarin-acrylate hybrids, whose fluorescence spectra have been measured.

Aβ Decreases Neprilysin-Mediated Alzheimer's Amyloid-β Peptide Degradation.

Alzheimer's disease is associated with the deposition of extracellular senile plaques, made primarily of amyloid-β (Aβ), particularly peptides Aβ and Aβ. Neprilysin, or neutral endopeptidase (NEP), catalyzes proteolysis of the amyloid peptides (Aβ) and is recognized as one of the major regulators of the levels of these peptides in the brain, preventing Aβ accumulation and plaque formation. Here, we used a combination of techniques to elucidate the mechanism of Aβ binding and cleavage by NEP.

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

The interaction of multi-LacNAc (Galβ1-4GlcNAc)-containing -(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.

Lipid transfer proteins and instructive regulation of lipid kinase activities: Implications for inositol lipid signaling and disease.

Cellular membranes are critical platforms for intracellular signaling that involve complex interfaces between lipids and proteins, and a web of interactions between a multitude of lipid metabolic pathways. Membrane lipids impart structural and functional information in this regulatory circuit that encompass biophysical parameters such as membrane thickness and fluidity, as well as chaperoning the interactions of protein binding partners. Phosphatidylinositol and its phosphorylated derivatives, the phosphoinositides, play key roles in intracellular membrane signaling, and these involvements are translated into an impressively diverse set of biological outcomes.

Functional diversification of the chemical landscapes of yeast Sec14-like phosphatidylinositol transfer protein lipid-binding cavities.

Phosphatidylinositol-transfer proteins (PITPs) are key regulators of lipid signaling in eukaryotic cells. These proteins both potentiate the activities of phosphatidylinositol (PtdIns) 4-OH kinases and help channel production of specific pools of phosphatidylinositol 4-phosphate (PtdIns(4)P) dedicated to specific biological outcomes. In this manner, PITPs represent a major contributor to the mechanisms by which the biological outcomes of phosphoinositide are diversified.

Fast and slow biomembrane solubilizing detergents: Insights into their mechanism of action.

Detergents are water-soluble amphiphiles. Above a critical concentration they self-organize in micelles and in the presence of phospholipids mixed micelles are formed. Much information is available on the structure of these self-assemblies and on the thermodynamics of their formation.

An equal opportunity collaboration between lipid metabolism and proteins in the control of membrane trafficking in the trans-Golgi and endosomal systems.

Recent years have witnessed the evolution of the cell biology of lipids into an extremely active area of investigation. Deciphering the involvement of lipid metabolism and lipid signaling in membrane trafficking pathways defines a major nexus of contemporary experimental activity on this front. Significant effort in that direction is invested in understanding the trans-Golgi network/endosomal system where unambiguous connections between membrane trafficking and inositol lipid and phosphatidylcholine metabolism were first discovered.

Polyamine-RNA-membrane interactions: From the past to the future in biology.

Biogenic polyamines (PAs), spermine, spermidine and putrescine are widely spread amino acid derivatives, present in living cells throughout the whole evolutionary scale. Their amino groups confer them a marked basic character at the cellular pH. We have tested the interaction of PAs with negatively-charged phospholipids in the absence and presence of nucleic acids (tRNA was mainly used for practical reasons).

Vesicular PtdIns(3,4,5)P3 and Rab7 are key effectors of sea urchin zygote nuclear membrane fusion.

Regulation of nuclear envelope dynamics is an important example of the universal phenomena of membrane fusion. The signalling molecules involved in nuclear membrane fusion might also be conserved during the formation of both pronuclear and zygote nuclear envelopes in the fertilised egg. Here, we determine that class-I phosphoinositide 3-kinases (PI3Ks) are needed for in vitro nuclear envelope formation.

Lipidic nanovesicles stabilize suspensions of metal oxide nanoparticles.

We have studied the effect of adding lipid nanovesicles (liposomes) on the aggregation of commercial titanium oxide (TiO2), zinc oxide (ZnO), or cerium oxide (CeO2) nanoparticles (NPs) suspensions in Hepes buffer. Liposomes were prepared with pure phospholipids or mixtures of phospholipids and/or cholesterol. Changes in turbidity were recorded as a function of time, either of metal nanoparticles alone, or for a mixture of nanoparticles and lipidic nanovesicles.

Endomembrane PtdIns(3,4,5)P3 activates the PI3K-Akt pathway.

PKB/Akt activation is a common step in tumour growth, proliferation and survival. Akt activation is understood to occur at the plasma membrane of cells in response to growth factor stimulation and local production of the phosphoinositide lipid phosphatidylinositol (3,4,5)-trisphosphate [PtdIns(3,4,5)P3] following phosphoinositide 3-kinase (PI3K) activation. The metabolism and turnover of phosphoinositides is complex--they act as signalling molecules as well as structural components of biological membranes.

Sec14-nodulin proteins and the patterning of phosphoinositide landmarks for developmental control of membrane morphogenesis.

Polarized membrane morphogenesis is a fundamental activity of eukaryotic cells. This process is essential for the biology of cells and tissues, and its execution demands exquisite temporal coordination of functionally diverse membrane signaling reactions with high spatial resolution. Moreover, mechanisms must exist to establish and preserve such organization in the face of randomizing forces that would diffuse it.

Histones cause aggregation and fusion of lipid vesicles containing phosphatidylinositol-4-phosphate.

In a previous article, we demonstrated that histones (H1 or histone octamers) interact with negatively charged bilayers and induce extensive aggregation of vesicles containing phosphatidylinositol-4-phosphate (PIP) and, to a lesser extent, vesicles containing phosphatidylinositol (PI). Here, we found that vesicles containing PIP, but not those containing PI, can undergo fusion induced by histones. Fusion was demonstrated through the observation of intervesicular mixing of total lipids and inner monolayer lipids, and by ultrastructural and confocal microscopy studies.

Histones and DNA compete for binding polyphosphoinositides in bilayers.

Recent discoveries on the presence and location of phosphoinositides in the eukaryotic cell nucleoplasm and nuclear membrane prompted us to study the putative interaction of chromatin components with these lipids in model membranes (liposomes). Turbidimetric studies revealed that a variety of histones and histone combinations (H1, H2AH2B, H3H4, octamers) caused a dose-dependent aggregation of phosphatidylcholine vesicles (large unilamellar vesicle or small unilamellar vesicle) containing negatively charged phospholipids. 5 mol % phosphatidylinositol-4-phosphate (PIP) was enough to cause extensive aggregation under our conditions, whereas with phosphatidylinositol (PI) at least 20 mol % was necessary to obtain a similar effect.