Biology subjects

Fragneto, G.

Publications and source records attributed to Fragneto, G..

4 recordsLinked to original sources

bioRxiv2025

Glycation enhances protein association with lipid bilayer membranes

Glycation is a non-enzymatic post-translational modification that leads to the formation of advanced glycation end-products (AGEs), which accumulate in the blood-stream under chronic hyperglycemia and are implicated in diabetes-related pathologies. While glycated proteins such as albumin or hemoglobin are widely used as biomarkers for glycemic control, the structural and chemical changes induced by glycation may also alter their interactions with lipid interfaces, including cellular membranes and lipoproteins, potentially affecting their biological distribution and diagnostic detectability. In this study, we investigated how glycation influences the interaction of bovine serum albumin (BSA) with supported lipid bilayers (SLBs) of different compositions, used as model systems to replicate the diversity of membrane surface charges and fluidity. Using neutron reflectometry (NR), we compared the membrane association of BSA and a chemically-enhanced glycated form of BSA (gBSA), focusing on nanostructural changes at the bilayer interface. Our results showed negligible interaction of either proteins with zwitterionic or cationic membranes. In contrast, both BSA and gBSA exhibited significant binding to negatively charged bilayers, with glycation significantly amplifying this interaction. Quantitatively, the membrane-associated protein volume fraction increased from 0.11 (BSA) to 0.17 (gBSA), suggesting that glycation modifies the proteins surface properties in ways that promote stronger lipid interactions with negatively charged membranes. These findings suggest that glycation not only affects protein structure but also modulates protein-membrane affinity in a lipid-dependent manner. This has important implications for the bioavailability and behavior of glycated albumin in the bloodstream, potentially influencing the accuracy of clinical assays and contributing to membrane-related pathophysiology in diabetes. Our work highlights the need for a deeper understanding of glycation-induced changes in protein-lipid interactions and their consequences for biomarker reliability and disease mechanisms. Graphical TOC Entry O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=82 SRC="FIGDIR/small/685514v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@150e71aorg.highwire.dtl.DTLVardef@749b35org.highwire.dtl.DTLVardef@179f468org.highwire.dtl.DTLVardef@19cfe8e_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2024

Untangling structural molecular details of the endocytic adaptor protein CALM upon binding with phosphatidylinositol 4,5-bisphosphate-containing model membranes

Clathrin assembly lymphoid myeloid leukemia protein (CALM) is involved in the formation of clathrin-mediated endocytic coats by virtue of binding many proteins involved in the process, including clathrin itself and AP2 cargo adaptor complex. CALM is able to specifically recognize the inner leaflet of the plasma membrane by binding the membranes phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2). Here, a biophysical approach, primarily using neutron and X-ray scattering and solid-state NMR experiments, was exploited to investigate CALM interaction with PtdIns(4,5)P2-presenting model membranes. The presented experimental data reveal how the CALM folded domain is partly accommodated within the lipid membrane, directly interacting with PtdIns(4,5)P2 phosphates. Moreover, these data suggest that CALMs amphiphilic N-terminal helix buries into the membrane, not only stabilising the protein docking to the membrane but also providing a mechanism to induce membrane curvature.

biophysics↗

bioRxiv2021

Strikingly different roles of SARS-CoV-2 fusion peptides uncovered by neutron scattering

Coronavirus disease-2019 (COVID-19), a lethal respiratory illness caused by the coronavirus SARS-CoV-2, emerged in the end of 2019 and has since spread aggressively across the globe. A thorough understanding of the molecular mechanisms of cellular infection by coronaviruses is therefore of utmost importance. A critical stage in infection is the fusion between viral and host membranes. Here, we present a detailed investigation of the role of the SARS-CoV-2 Spike protein, and the influence of calcium and cholesterol, in this fusion process. Structural information from specular neutron reflectometry and small angle neutron scattering, complemented by dynamics information from quasi-elastic and spin-echo neutron spectroscopy, revealed strikingly different functions encoded in the Spike fusion domain. Calcium drives the N-terminal of the Spike fusion domain to fully cross the host plasma membrane. Removing calcium however re-orients the protein to the lipid leaflet in contact with the virus, leading to significant changes in lipid fluidity and rigidity. In conjunction with other regions of the fusion domain which are also positioned to bridge and dehydrate viral and host membranes, the molecular events leading to cell entry by SARS-CoV-2 are proposed. SIGNIFICANCEWe have recreated here important elements of the critical membrane fusion mechanism of the SARS-CoV-2 coronavirus by simplifying the system down to its core elements, amenable to experimental analysis by neutron scattering. Neutrons are well suited for the study of protein - membrane interactions under physiological conditions, since they allow structural and dynamics characterization at room temperature. Our results revealed strikingly different functions encoded in the viral Spike fusion domain and thereby provide a potential calcium-dependent cell entry mechanism for SARS-CoV-2. In particular, calcium drives the proteins N-terminal to harpoon through the host membrane, while removing calcium re-orients the protein so that it is able to bridge and dehydrate lipid membranes, facilitating their fusion.

biophysics↗

bioRxiv2020

Purification, characterization and influence on membrane properties of the plant-specific sphingolipids GIPC

The plant plasma membrane (PM) is an essential barrier between the cell and the external environment. The PM is crucial for signal perception and transmission. It consists of an asymmetrical lipid bilayer made up of three different lipid classes: sphingolipids, sterols and phospholipids. The most abundant sphingolipids in the plant PM are the Glycosyl Inositol Phosphoryl Ceramides (GIPCs), representing up to 40% of total sphingolipids, assumed to be almost exclusively in the outer leaflet of the PM. In this study, we investigated the structure of GIPCs and their role in membrane organization. Since GIPCs are not commercially available, we developed a protocol to extract and isolate GIPC-enriched fractions from eudicots (cauliflower and tobacco) and monocots (leek and rice). Lipidomic analysis confirmed the presence of different long chain bases and fatty acids. The glycan head groups of the different GIPC series from monocots and dicots were analysed by GC-MS showing different sugar moieties. Multiple biophysics tools namely Langmuir monolayer, {zeta}-Potential, light scattering, neutron reflectivity, solid state 2H-NMR and molecular modelling were used to investigate the physical properties of the GIPCs, as well as their interaction with free and conjugated phytosterols. We showed that GIPCs increase the thickness and electronegativity of model membranes, interact differentially with the phytosterols species and regulate the gel-to-fluid phase transition during temperature variations.

plant biology↗