Key carboxylate residues for iron transit through the prokaryotic ferritin SynFtn

Ferritins are proteins forming 24meric rhombic dodecahedral cages that play a key role in iron storage and detoxification in all cell types. Their function requires the transport of Fe2+ from the exterior of the protein to buried di-iron catalytic sites, known as ferroxidase centres, where Fe2+ is oxidized to form Fe3+-oxo precursors of the ferritin mineral core. The route of iron transit through animal ferritins is well understood: the Fe2+ substrate enters the protein via channels at the threefold axes and conserved carboxylates on the inner surface of the protein cage have been shown to contribute to transient binding sites that guide Fe2+ to the ferroxidase centres. The routes of iron transit through prokaryotic ferritins are less well studied but for some, at least, there is evidence that channels at the twofold axes are the major route for Fe2+ uptake. SynFtn, isolated from the cyanobacterium Synechococcus CC9311, is an atypical prokaryotic ferritin that was recently shown to take up Fe2+ via its threefold channels. However, the transfer site carboxylate residues conserved in animal ferritins are absent, meaning that the route taken from the site of iron entry into SynFtn to the catalytic centre is yet to be defined. Here, we report the use of a combination of site-directed mutagenesis, absorbance-monitored activity assays and protein crystallography to probe the effect of substitution of two residues potentially involved in this pathway. Both Glu141 and Asp65 play a role in guiding the Fe2+ substrate to the ferroxidase centre. In the absence of Asp65, routes for Fe2+ to, and Fe3+ exit from, the ferroxidase centre are affected resulting in inefficient formation of the mineral core. These observations further define the iron transit route in what may be the first characterized example of a new class of ferritins peculiar to cyanobacteria.

Longitudinal and Transverse Relaxivity Analysis of Native Ferritin and Magnetoferritin at 7 T MRI

Magnetite mineralization in human tissue is associated with various pathological processes, especially neurodegenerative disorders. Ferritin’s mineral core is believed to be a precursor of magnetite mineralization. Magnetoferritin (MF) was prepared with different iron loading factors (LFs) as a model system for pathological ferritin to analyze its MRI relaxivity properties compared to those of native ferritin (NF). The results revealed that MF differs statistically significantly from NF, with the same LF, for all studied relaxation parameters at 7 T: r1, r2, r2*, r2/r1, r2*/r1. Distinguishability of MF from NF may be useful in non-invasive MRI diagnosis of pathological processes associated with iron accumulation and magnetite mineralization (e.g., neurodegenerative disorders, cancer, and diseases of the heart, lung and liver). In addition, it was found that MF samples possess very strong correlation and MF’s relaxivity is linearly dependent on the LF, and the transverse and longitudinal ratios r2/r1 and r2*/r1 possess complementary information. This is useful in eliminating false-positive hypointensive artefacts and diagnosis of the different stages of pathology. These findings could contribute to the exploitation of MRI techniques in the non-invasive diagnosis of iron-related pathological processes in human tissue.

Quantification of Iron Release from Native Ferritin and Magnetoferritin Induced by Vitamins B2 and C

Various pathological processes in humans are associated with biogenic iron accumulation and the mineralization of iron oxide nanoparticles, especially magnetite. Ferritin has been proposed as a precursor to pathological magnetite mineralization. This study quantifies spectroscopically the release of ferrous ions from native ferritin and magnetoferritin as a model system for pathological ferritin in the presence of potent natural reducing agents (vitamins C and B2) over time. Ferrous cations are required for the transformation of ferrihydrite (physiological) into a magnetite (pathological) mineral core and are considered toxic at elevated levels. The study shows a significant difference in the reduction and iron release from native ferritin compared to magnetoferritin for both vitamins. The amount of reduced iron formed from a magnetoferritin mineral core is two to five times higher than from native ferritin. Surprisingly, increasing the concentration of the reducing agent affects only iron release from native ferritin. Magnetoferritin cores with different loading factors seem to be insensitive to different concentrations of vitamins. An alternative hypothesis of human tissue magnetite mineralization and the process of iron-induced pathology is proposed. The results could contribute to evidence of the molecular mechanisms of various iron-related pathologies, including neurodegeneration.

STUDY OF PHYSICAL-MECHANICAL PROPERTIES OF ASPHALT-CONCRETE SPECIMENS ON ACTIVATED MINERAL POWDERS OF VARIOUS COMPOSITIONS

the article presents the results of testing asphalt samples based on activated mineral powder from silica-containing raw materials. Activated filler was obtained by joint grinding of waste wet magnetic separation (WMS) of ferruginous quartzites of the Kursk Magnetic Anomaly in a spherical planetary mill in the presence of surfactants (stearic acid, adhesive additive Amdor-10) and bitumen. First of all, it should be noted that the use of activated mineral powder in the composition of asphalt concrete mixture led to an increase in the packing density of the composite. This was reflected not only in the technological process, but also contributed to the decrease in the porosity of the mineral core. It was found that the use of surfactants in the process of grinding mineral powder made it possible to increase the strength of asphalt concrete samples. However, the change in the crack resistance and shear resistance of asphalt concrete specimens, determined from the ratio of the corresponding compressive strengths, did not occur (table). This is due to the fact that there has been a proportional change in all the strength characteristics at different temperatures. Analysis of the physical-mechanical characteristics of the properties of asphalt concrete led to the conclusion that it was possible to use activated mineral powder from waste WMS as a component of asphalt concrete mixtures.

Emerging and Dynamic Biomedical Uses of Ferritin

Ferritin, a ubiquitously expressed protein, has classically been considered the main iron cellular storage molecule in the body. Owing to the ferroxidase activity of the H-subunit and the nucleation ability of the L-subunit, ferritin can store a large amount of iron within its mineral core. However, recent evidence has demonstrated a range of abilities of ferritin that extends well beyond the scope of iron storage. This review aims to discuss novel functions and biomedical uses of ferritin in the processes of iron delivery, delivery of biologics such as chemotherapies and contrast agents, and the utility of ferritin as a biomarker in a number of neurological diseases.

Reductive Mobilization of Iron from Intact Ferritin: Mechanisms and Physiological Implication

Ferritins are highly conserved supramolecular protein nanostructures composed of two different subunit types, H (heavy) and L (light). The two subunits co-assemble into a 24-subunit heteropolymer, with tissue specific distributions, to form shell-like protein structures within which thousands of iron atoms are stored as a soluble inorganic ferric iron core. In-vitro (or in cell free systems), the mechanisms of iron(II) oxidation and formation of the mineral core have been extensively investigated, although it is still unclear how iron is loaded into the protein in-vivo. In contrast, there is a wide spread belief that the major pathway of iron mobilization from ferritin involves a lysosomal proteolytic degradation of ferritin, and the dissolution of the iron mineral core. However, it is still unclear whether other auxiliary iron mobilization mechanisms, involving physiological reducing agents and/or cellular reductases, contribute to the release of iron from ferritin. In vitro iron mobilization from ferritin can be achieved using different reducing agents, capable of easily reducing the ferritin iron core, to produce soluble ferrous ions that are subsequently chelated by strong iron(II)-chelating agents. Here, we review our current understanding of iron mobilization from ferritin by various reducing agents, and report on recent results from our laboratory, in support of a mechanism that involves a one-electron transfer through the protein shell to the iron mineral core. The physiological significance of the iron reductive mobilization from ferritin by the non-enzymatic FMN/NAD(P)H system is also discussed.

On the mineral core of ferritin-like proteins: structural and magnetic characterization

We report a combined structural and magnetic study of the mineral core biomineralized by horse spleen ferritin and three prokaryotic ferritin-like proteins: bacterial ferritin and bacterioferritin from Escherichia coli and archaeal ferritin from Pyrococcus furiosus.