Daniela Russo

Due to its unique chemical structure, lycopene may induce multiple health benefits as a natural antioxidant. However, the biological properties of lycopene and, therefore, its wide applications in foods and feed, food supplements, pharmaceutics and cosmetics are limited due to its specific hydrophobic nature and low chemical stability. The specific biological and physicochemical characteristics are due to the presence of 11 conjugated double bonds, which highlights it among the >700 carotenoids discovered, being valued as the most efficient singlet oxygen quencher. However, due to its highly unsaturated hydrocarbon structure, lycopene is highly susceptible to chemical degradation when exposed to high temperatures, oxygen, and light, leading to color fading and the loss of bioactive properties. The stability, bioaccessibility and bioavailability of lycopene may be improved by using encapsulation, such as spray and freeze-drying, emulsions, liposomes, complex coacervation, electrospinning, etc. The main purpose of this study was to highlight the main characteristics of lycopene, the main sources, the principle and key factors of encapsulation, thus covering the most used and effective stabilization methods described in the literature. An important issue covered by this work is related to the bioaccessibility and bioavailability of different isomers of lycopene, with the main purpose of clarifying the mechanism of digestion.

This study investigates the structural dynamics of the TRAF2 C-terminal domain (TRAF2-C), a key adaptor protein in TNF receptor signaling. TRAF2 usually forms trimers, but its ability to dissociate into monomers is critical for regulating apoptosis, inflammation, and cell survival. Using Fluorescence Fluctuation Spectroscopy, dynamic light scattering, circular dichroism, and Small Angle Neutron Scattering, we analyzed TRAF2-C over a wide concentration range. At nanomolar levels, the protein dissociates easily, with trimers representing only a minor fraction, while micromolar concentrations strongly favor trimerization. Dissociation also reduces α-helical content without disrupting the overall fold. Molecular dynamics simulations and protein contact network analysis support this analysis, identifying interfacial residues and hydrogen bonds as key factors stabilizing oligomers and enabling dynamic asymmetry. Overall, these findings highlight TRAF2-C’s capacity to switch between monomeric and trimeric states as a crucial regulatory mechanism, offering insights into TRAF-mediated signaling and potential therapeutic strategies.

Protein–polymer conjugates are engineered to enhance the pharmacokinetics and stability of biopharmaceuticals. This review delves into the structural and dynamic characteristics of protein–polymer conjugates, employing advanced techniques such as neutron scattering, circular dichroism, and fluorescence spectroscopy. Here, we focus on model proteins like maltose‐binding protein (MBP), bovine serum albumin (BSA), and myoglobin (Mb) conjugated with hydrolysable polyphosphoesters (PPEs) to generate fully degradable polymer–protein conjugates. The study underscores the influence of factors such as the polymers' molar mass, grafting density, hydration levels, and deuteration on protein stability, flexibility, and thermal properties. The study demonstrates that the degree of polymerization, solvation properties, and isotopic composition play crucial roles in determining the behavior of protein–polymer complexes. In particular, neutron scattering techniques are invaluable in unraveling the interaction mechanisms, providing valuable insights that can inform the optimization of protein–polymer conjugates for therapeutic applications.

The conjugation of proteins with polymers offers immense biotechnological potential by creating novel macromolecules. This article presents experimental findings on the structural properties of maltose‐binding protein (MBP) conjugated with linear biodegradable polyphosphoester polymers with different molecular weights. We studied isotopic effects on both proteins and polymers. Circular dichroism and fluorescence spectroscopy and small‐angle neutron scattering reveal that the conjugation process destabilizes the protein, affecting the secondary more than the tertiary structure, even at room temperature, and that the presence of two domains in the MBP may contribute to its observed instability. Notably, unfolding temperatures differ between native MBP and the conjugates. In particular, this study sheds light on the complex interplay of factors such as the deuteration influencing protein stability and conformational changes in the conjugation processes. The perdeuteration influences the hydrogen bond network and hydrophobic interactions in the case of the MBP protein. The perdeuteration of the protein influences the hydrogen bond network and hydrophobic interactions. This is evident in the decreased thermal stability of deuterated MBP protein, in the conjugate, especially with high‐molecular‐mass polymers.

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. SIGNIFICANCE We 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 protein’s 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.

One can take advantage of the influence of a polymer conjugated with a protein to control the thermal stability and the deployment of the protein. Here, the structural properties are reported of the protein–polymer conjugate myoglobin (Mb)‐poly(ethyl ethylene phosphate) (PEEP) in the native and unfolded conformations, in order to understand the respective roles of the protein and of the polymer size in the stability of the conjugate. The effect is also investigated of the grafting density of the linear biodegradable polyphosphoesters covalently attached to the protein. It is observed that, while the conjugation process at room temperature does not modify the secondary and tertiary structure of the Mb, the unfolding process, as a function of temperature, depends on the grafting density. Small angle neutron scattering reveals that, at room temperature, conjugation does not alter the size of the native protein and that the thickness of the polymer shell around the protein increases as a function of grafting density and of polymer molecular weight. The denatured form of all conjugates is described by an unfolded chain and a correlation length due to the presence of local stiffness.

The bio and chemical physics of protein–polymer conjugates are related to parameters that characterize each component. With this work, it is intended to feature the dynamical properties of the protein–polymer conjugate myoglobin (Mb)–poly(ethyl ethylene phosphate), in the ps and ns time scales, in order to understand the respective roles of the protein and of the polymer size in the dynamics of the conjugate. Elastic and quasi‐elastic neutron scattering is performed on completely hydrogenated samples with variable number of polymer chains covalently attached to the protein. The role of the polymer length in the protein solvation and internal dynamics is investigated using two conjugates formed by polymers of different molecular weight. It is confirmed that the flexibility of the complex increases with the number of grafted polymer chains and that a sharp dynamical transition appears when either grafting density or polymer molecular weight are high. It is shown that protein size is crucial for the polymer structural organization and interaction on the protein surface and it is established that the glass properties of the polymer change upon conjugation. The results give a better insight of the equivalence of the polymer coating and the role of water on the surface of proteins.

Many biomedical applications employ covalent attachment to synthetic polymers to enhance the efficiency of proteins or other therapeutically active molecules. We report here the impact of polymer conjugation on the structural and thermal stability of a protein model, the bovine serum albumin, using a variable number of linear biodegradable polyphosphoesters, which were covalently tethered to the protein. We observed that BSAs’ secondary structure measured by circular dichroism is independent of the conjugation. Small angle neutron scattering, however reveals a change from ellipsoid to globular shape of the whole complex arising from a slight compaction of the protein core and an increase of the polymer’s radius of gyration as a function of the grafting polymer density. In particular, we highlight a gradual change of the polymer conformation around the protein and elongation of the semi-major dimension of the ellipsoidal protein. Our results will contribute to the description of biophysical characteristics of a new class of biologically relevant protein-polymer conjugates

We investigate the relaxation dynamics of protein-polymer conjugates by neutron scattering spectroscopy to understand to which extent the coating of a protein by a polymer can replace water in promoting thermal structural fluctuations. For this purpose, we compare the dynamics of protein - polymer mixtures to conjugates with variable number of polymers covalently attached to the protein. Results show that the flexibility of the protein is larger in protein - polymer mixtures than in native protein or in conjugates, even in the dry state. Upon hydration, both the native protein and the conjugate show an equivalent relaxation dynamics suggesting that the polymer at the protein surface adsorbs all water molecules.