Destruction of Lysozyme Amyloid Fibrils Induced by Magnetoferritin and Reconstructed Ferritin.

Neurodegenerative disorders, including Alzheimer's disease (AD), Parkinson's disease (PD), or systemic amyloidosis, are characterized by the specific protein transformation from the native state to stable insoluble deposits, e.g., amyloid plaques. The design of potential therapeutic agents and drugs focuses on the destabilization of the bonds in their beta-rich structures. Surprisingly, ferritin derivatives have recently been proposed to destabilize fibril structures. Using atomic force microscopy (AFM) and fluorescence spectrophotometry, we confirmed the destructive effect of reconstructed ferritin (RF) and magnetoferritin (MF) on lysosome amyloid fibrils (LAF). The presence of iron was shown to be the main factor responsible for the destruction of LAF. Moreover, we found that the interaction of RF and MF with LAF caused a significant increase in the release of potentially harmful ferrous ions. Zeta potential and UV spectroscopic measurements of LAF and ferritin derivative mixtures revealed a considerable difference in RF compared to MF. Our results contribute to a better understanding of the mechanism of fibril destabilization by ferritin-like proteins. From this point of view, ferritin derivatives seem to have a dual effect: therapeutic (fibril destruction) and adverse (oxidative stress initiated by increased Fe2+ release). Thus, ferritins may play a significant role in various future biomedical applications.

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.

The Impact of Redox, Hydrolysis and Dehydration Chemistry on the Structural and Magnetic Properties of Magnetoferritin Prepared in Variable Thermal Conditions.

Ferritin, a spherically shaped protein complex, is responsible for iron storage in bacteria, plants, animals, and humans. Various ferritin iron core compositions in organisms are associated with specific living requirements, health state, and different biochemical roles of ferritin isomers. Magnetoferritin, a synthetic ferritin derivative, serves as an artificial model system of unusual iron phase structures found in humans. We present the results of a complex structural study of magnetoferritins prepared by controlled in vitro synthesis. Using various complementary methods, it was observed that manipulation of the synthesis technology can improve the physicochemical parameters of the system, which is useful in applications. Thus, a higher synthesis temperature leads to an increase in magnetization due to the formation of the magnetite phase. An increase in the iron loading factor has a more pronounced impact on the protein shell structure in comparison with the pH of the aqueous medium. On the other hand, a higher loading factor at physiological temperature enhances the formation of an amorphous phase instead of magnetite crystallization. It was confirmed that the iron-overloading effect alone (observed during pathological events) cannot contribute to the formation of magnetite.

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.

Fe(II) formation after interaction of the amyloid ß-peptide with iron-storage protein ferritin.

The interaction of amyloid ß-peptide (Aß) with the iron-storage protein ferritin was studied in vitro. We have shown that Aß during fibril formation process is able to reduce Fe(III) from the ferritin core (ferrihydrite) to Fe(II). The Aß-mediated Fe(III) reduction yielded a two-times-higher concentration of free Fe(II) than the spontaneous formation of Fe(II) by the ferritin itself. We suggest that Aß can also act as a ferritin-specific metallochaperone-like molecule capturing Fe(III) from the ferritin ferrihydrite core. Our observation may partially explain the formation of Fe(II)-containing minerals in human brains suffering by neurodegenerative diseases.

Dechlorination of 2,4,4'-trichlorobiphenyl by magnetoferritin with different loading factors.

Polychlorinated biphenyls are synthetic industrial organic substances. These persistent pollutants occur in nature causing high ecological risks and damage to human health. Magnetoferritin nanoparticles composed of apoferritin protein shell surrounding synthetically prepared iron-based nanoparticles seem to be a promising candidate for polychlorinated biphenyls elimination. Properties of magnetoferritin, as a redox activity, a biocompatible character, high application possibilities and a close relationship with the human body promoted ours in vitro investigation of the magnetoferritin catalytic activity in the presence of representative 2,4,4'-trichlorobiphenyl. Basic physico-chemical properties of magnetoferritin were determined by ultraviolet and visible spectrophotometry, dynamic light scattering, zeta potential measurements, superconducting quantum interference device magnetometry and atomic force microscopy. The remediation effect of magnetoferritin on 2,4,4'-trichlorobiphenyl was demonstrated by the use of gas chromatography in combination with infrared spectroscopy.

Morphology and Magnetic Structure of the Ferritin Core during Iron Loading and Release by Magnetooptical and NMR Methods.

Ferritins are proteins, which serve as a storage and transportation capsule for iron inside living organisms. Continuously charging the proteins with iron and releasing it from the ferritin is necessary to assure proper management of these important ions within the organism. On the other hand, synthetic ferritins have great potential for biomedical and technological applications. In this work, the behavior of ferritin during the processes of iron loading and release was examined using multiplicity of the experimental technique. The quality of the protein's shell was monitored using circular dichroism, whereas the average size and its distribution were estimated from dynamic light scattering and transmission electron microscopy images, respectively. Because of the magnetic behavior of the iron mineral, a number of magnetooptical methods were used to gain information on the iron core of the ferritin. Faraday rotation and magnetic linear birefringence studies provide evidence that the iron loading and the iron-release processes are not symmetrical. The spatial organization of the mineral within the protein's core changes depending on whether the iron was incorporated into or removed from the ferritin's shell. Magnetic optical rotatory dispersion spectra exclude the contribution of the Fe(II)-composed mineral, whereas joined magnetooptical and nuclear magnetic resonance results indicate that no mineral with high magnetization appear at any stage of the loading/release process. These findings suggest that the iron core of loaded/released ferritin consists of single-phase, that is, ferrihydrite. The presented results demonstrate the usefulness of emerging magnetooptical methods in biomedical research and applications.