Hydrogen peroxide induces heme degradation and protein aggregation in human neuroglobin: roles of the disulfide bridge and hydrogen-bonding in the distal heme cavity.

In the present study, human neuroglobin (hNgb) was found to undergo H2 O2 -induced breakdown of the heme center at a much slower rate than other globins, namely in the timescale of hours against minutes. We investigated how the rate of the process is affected by the Cys46/Cys55 disulfide bond and the network of non-covalent interactions in the distal heme side involving Tyr44, Lys67, the His64 heme iron axial ligand and the heme propionate-7. The rate is increased by the Tyr44 to Ala and Phe mutations; however the rate is lowered by Lys67 to Ala swapping. The absence of the disulfide bridge slows down the reaction further. Therefore, the disulfide bond-controlled accessibility of the heme site and the residues at position 44 and 67 affect the activation barrier of the reaction. Wild-type and mutated species form ß-amyloid aggregates in the presence of H2 O2 producing globular structures. Furthermore, the C46A/C55A, Y44A, Y44F and Y44F/C46A/C55A variants yield potentially harmful fibrils. Finally, the nucleation and growth kinetics for the aggregation of the amyloid structures can be successfully described by the Finke-Watzky model.

Al-Substituted Tobermorites: An Effective Cation Exchanger Synthesized from "End-of-Waste" Materials.

The policies to meet the "zero waste" regime and transition to sustainable circular economy can no longer ignore the use of wastes in place of natural resources, and these daunting and vital societal challenges are nowadays being faced by several nations. The main objective of this work was to search waste materials suitable for a quick and environmentally friendly production of a nanoporous geomaterial able to trap toxic metals at the solid/liquid interface. More specifically, the end-of-waste from the thermal inertization of cement-asbestos and glass powder from domestic glass containers have been employed as sources for the hydrothermal synthesis of a tobermorite-rich material (TRM) successfully tested for the selective removal of Pb2+, Zn2+, Cd2+, and Ni2+ from aqueous solutions. The synthesis was carried out in alkaline solution under mild hydrothermal conditions (120 °C) within 24 h. The quantitative phase analyses of the TRM carried out by applying the Rietveld method showed the occurrence of a large amount of well-crystallized 11 Å Al-substituted tobermorites and an amorphous phase and a lower content of aragonite and calcite. Chemical analyses and thermogravimetric measurements coupled with simultaneous evolved gas mass spectrometry highlighted that Al3+ for Si4+ substitutions in the wollastonite-like tetrahedral chains of tobermorites are balanced by the occurrence of Ca2+, Na+, and K+ cations in the interlayer rather than by (OH)- for O2- substitutions in the CaO polyhedra. Time-dependent removal tests clearly indicated that metal cations are selectively adsorbed depending on their concentration in solution. Moreover, the kinetic curves showed that the removal of metals from solution is fast and the equilibrium is almost reached in the first 8 h.

pH-Dependent peroxidase activity of yeast cytochrome c and its triple mutant adsorbed on kaolinite.

The peroxidase activity of wild-type yeast cytochrome c and its triple mutant K72AK73AK79A adsorbed onto kaolinite was investigated as a function of pH and temperature. Both adsorbed proteins displayed an appreciable catalytic activity, which remained constant from pH 7 to pH 10, decreased below pH 7, and showed a remarkable increase at pH values lower than 4. In the whole pH range investigated the catalytic activity of the adsorbed wild-type cytochrome c was higher than that of the mutant. Both diffuse-reflectance UV-vis and resonance Raman spectroscopies applied on solid samples were used to probe the structural features responsible for the catalytic activity of the immobilized proteins. At neutral and alkaline pH values a six-coordinate low-spin form of cytochrome c was observed, while at pH < 7 the formation of a high-spin species occurred whose population increased at decreasing pH. The orientation and exposure of the heme to the substrate-strictly dependent on adsorption-was found to affect the peroxidase activity.

Gaseous Heptanethiol Removal by a Fe3+-Phenanthroline-Kaolinite Hybrid Material.

Kaolinite functionalized by the µ-oxo Fe3+-phenanthroline complex (Fe+3Phen) was selected to test its ability to efficiently remove and store gaseous heptanethiol (HPT). Spectroscopic techniques, elemental analysis, and thermal analysis coupled with evolved gas mass spectrometry were employed to characterize the material before and after the exposure to the gas and to define the adsorption process. The amount of HPT trapped by the functionalized kaolinite after 60 days is 0.10940 moles per 100 g of kaolinite which, considering the amount of adsorbed Fe+3Phen (0.00114 moles per 100 g of kaolinite), means a thiol/Fe3+Phen molar ratio of about 100:1, a value much higher than those found in the past for Fe+3Phen functionalized montmorillonite and sepiolite. In addition, the process was found to be efficient also beyond 60 days. This significant removal of the smelly gas was explained by considering a continuous catalytic activity of Fe3+ toward the oxidation of thiol to disulfide.

Phosphorylated cofilin-2 is more prone to oxidative modifications on Cys39 and favors amyloid fibril formation.

Cofilins are small protein of the actin depolymerizing family. Actin polymerization/depolymerization is central to a number of critical cellular physiological tasks making cofilin a key protein for several physiological functions of the cell. Cofilin activity is mainly regulated by phosphorylation on serine residue 3 making this post-translational modification key to the regulation of myofilament integrity. In fact, in this form, the protein segregates in myocardial aggregates in human idiopathic dilated cardiomyopathy. Since myofilament network is an early target of oxidative stress we investigated the molecular changes induced by oxidation on cofilin isoforms and their interplay with the protein phosphorylation state to get insight on whether/how those changes may predispose to early protein aggregation. Using different and complementary approaches we characterized the aggregation properties of cofilin-2 and its phosphomimetic variant (S3D) in response to oxidative stress in silico, in vitro and on isolated cardiomyocytes. We found that the phosphorylated (inactive) form of cofilin-2 is mechanistically linked to the formation of an extended network of fibrillar structures induced by oxidative stress via the formation of a disulfide bond between Cys39 and Cys80. Such phosphorylation-dependent effect is likely controlled by changes in the hydrogen bonding network involving Cys39. We found that the sulfide ion inhibits the formation of such structures. This might represent the mechanism for the protective effect of the therapeutic agent Na2S on ischemic injury.

Electrochemical data on redox properties of human Cofilin-2 and its Mutant S3D.

The reported data are related to a research paper entitled "Phosphorylated cofilin-2 is more prone to oxidative modifications on Cys39 and favors amyloid fibril formation" [1]. Info about the formation and redox properties of the disulfide bridge of a protein is quite difficult to obtain and only in a few cases was it possible to observe a cyclic voltammetry (CV) signal [2,3]. Human cofilin-2 contains two cysteines (Cys39 and Cys80) which can be oxidized in suitable conditions and form a disulfide bridge [1]. For this purpose, CV measurements were carried out on human cofilin-2 WT and its mutant S3D immobilized on a gold electrode coated by an anionic self-assembled monolayer (SAM), after a pre-oxidation time which was fundamental for observing a CV signal relating to the oxidation/reduction process of the disulfide bridge of the proteins. The data include CV curves obtained with and without electrochemical pre-oxidation and after oxidation with H2O2. In addition, the plot of the cathodic peak current vs. electrochemical pre-oxidation time and the pH dependence of the formal potential (E°') are reported. The data obtained by CV measurements were used to determine the time required to form the disulfide bridge for the immobilized proteins and, consequently, to observe the CV signal, to calculate the E°' values and analyse the pH dependence of E°'. The electrochemical data were provided which will be useful for further electrochemical investigations regarding proteins bearing disulfide bridge(s) or cysteines prone to oxidation.

Trapping at the Solid-Gas Interface: Selective Adsorption of Naphthalene by Montmorillonite Intercalated with a Fe(III)-Phenanthroline Complex.

In this study, stable hybrid materials (Mt-Fe(III)Phen), made by the µ-oxo Fe(III)-phenanthroline complex [(OH2)3(Phen)FeOFe(Phen)(OH2)3]4+ (Fe(III)Phen) intercalated in different amounts into montmorillonite (Mt), were used as a trap for immobilizing gaseous benzene and naphthalene and their mono chloro-derivatives at 25 and 50 °C. The entrapping process was studied through elemental analysis, magic angle spinning NMR spectroscopy, thermal analysis, and evolved gas mass spectrometry. Naphthalene and 1-chloronaphthalene were found to be immobilized in large amount at both temperatures. Molecular modeling allowed designing of the structure of the interlayer in the presence of the immobilized aromatic molecules. Adsorption is affected by the amount of the Fe complex hosted in the interlayer of the entrapping hybrid materials. On the contrary, under the same conditions, benzene and chlorobenzene were not adsorbed. Thermal desorption of naphthalenes was obtained under mild conditions, and immobilization was found to be reversible at least for 20 adsorption/desorption cycles.

Effective and Selective Trapping of Volatile Organic Sulfur Derivatives by Montmorillonite Intercalated with a µ-oxo Fe(III)-Phenanthroline Complex.

The µ-oxo Fe(III)-phenanthroline complex [(OH2)3(Phen)FeOFe(Phen) (OH2)3]+4 intercalated in montmorillonite provides a stable hybrid material. In this study, the ability and efficiency of this material to immobilize thiols in gas phase, acting as a trap at the solid-gas interface, were investigated. Aliphatic thiols containing both hydrophilic and hydrophobic end groups were chosen to test the selectivity of this gas trap. DR-UV-vis, IR, elemental analysis, thermal analysis and evolved gas mass spectrometry, X-ray powder diffraction, and X-ray absorption spectroscopy techniques were employed to characterize the hybrid material before and after thiol exposure and to provide information on the entrapping process. Thiol immobilization is very large, up to 21% w/w for heptanethiol. In addition, evidence was obtained that immobilization occurs through the formation of a covalent bond between the iron of the complex and the sulfur of the thiol. This provides an immobilization process characterized by a higher stability with respect to the methods based on physi-adsorption. Thiol immobilization resulted thermally reversible at least for 20 adsorption/desorption cycles. Unlike standard desulfurization processes like hydrotreating and catalytic oxidation which work at high temperatures and pressures, the present system is able to efficiently trap thiols at room temperature and pressure, thus saving energy. Furthermore, we found that the selectivity of thiol immobilization can be tuned acting on the amount of complex intercalated in montmorillonite. In particular, montmorillonite semisaturated with the complex captures both hydrophobic and hydrophilic thiols, while the saturated montmorillonite shows a strong selectivity toward the hydrophobic molecules.

Pre-amyloid oligomers budding:a metastatic mechanism of proteotoxicity.

The pathological hallmark of misfolded protein diseases and aging is the accumulation of proteotoxic aggregates. However, the mechanisms of proteotoxicity and the dynamic changes in fiber formation and dissemination remain unclear, preventing a cure. Here we adopted a reductionist approach and used atomic force microscopy to define the temporal and spatial changes of amyloid aggregates, their modes of dissemination and the biochemical changes that may influence their growth. We show that pre-amyloid oligomers (PAO) mature to form linear and circular protofibrils, and amyloid fibers, and those can break reforming PAO that can migrate invading neighbor structures. Simulating the effect of immunotherapy modifies the dynamics of PAO formation. Anti-fibers as well as anti-PAO antibodies fragment the amyloid fibers, however the fragmentation using anti-fibers antibodies favored the migration of PAO. In conclusion, we provide evidence for the mechanisms of misfolded protein maturation and propagation and the effects of interventions on the resolution and dissemination of amyloid pathology.