Plant Ferritin Research Articles

The potential for the use of leghemoglobin and plant ferritin as sources of iron.

Iron is an essential component for the body, but it is also a major cause for the development of many diseases such as cancer, cardiovascular diseases, and autoimmune diseases. It has been suggested that a diet rich in meat products, especially red meat and highly processed products, constitute a nutritional model that increases the risk of developing. In this context, it is indicated that people on an elimination diet (vegetarians and vegans) may be at risk of deficiencies in iron, because this micronutrient is found mainly in foods of animal origin and has lower bioavailability in plant foods. This article reviews the knowledge on the use of leghemoglobin and plant ferritin as sources of iron and discusses their potential for use in vegetarian and vegan diets.

Cadmium and Zn hyperaccumulation provide efficient constitutive defense against Turnip yellow mosaic virus infection in Noccaea caerulescens

To understand the role of Zn and Cd in anti-viral defence, Zn/Cd hyperaccumulator Noccaea caerulescens plants grown with deficient (0.3 µM), replete (10 µM) and excess (100 µM) Zn2+ and Cd (10 µM Zn2+ + 1 µM Cd2+) were infected with Turnip yellow mosaic virus (TYMV). Gas exchange and chlorophyll fluorescence kinetics analyses demonstrated direct TYMV effects on photosynthetic light reactions but N. caerulescens was more resistant against TYMV than the previously studied non-hyperaccumulator N. ochroleucum. Virus abundance and photosynthesis inhibition were the lowest in the high Zn and Cd treatments. RNAseq analysis of 10 µM Zn2+ plants revealed TYMV-induced upregulation of Ca transporters, chloroplastic ZTP29 and defence genes, but none of those that are known to be strongly involved in hyperaccumulation. Synchrotron µ-XRF tomography, however, showed that Zn hyperaccumulation remained strongest in vacuoles of epidermal storage cells regardless of infection. This was in contrast to N. ochroleucum, where apoplastic Zn drastically increased in response to TYMV. These results suggest that the antiviral response of N. caerulescens is less induced by the onset of this biotic stress, but it is rather a permanent resistant state of the plant. Real-time qPCR revealed upregulation of ferritin in Zn10 infected plants, suggesting Fe deprivation as a virus defence strategy under suboptimal Zn supply.

A Dual Function of Ferritin (Animal and Plant): Its Holo Form for Iron Supplementation and Apo Form for Delivery Systems.

Ferritins represent a class of iron storage proteins with detoxification functions. The importance of these proteins is reflected by their wide distribution throughout the animal and plant kingdoms. Ferritin has two forms: holo and apo. Holo ferritin can act as an efficient and safe factor for iron supplementation, whereas apo ferritin is able to serve as a promising delivery nanovehicle for nutrients and bioactive compounds. So far, the dual functions of ferritins from animal and plant sources have been extensively studied in several fields, such as food, nutrition, medicine, and materials. This review outlines the structure of animal and plant ferritin, the iron supplementation function of holo ferritin, and the delivery function of apo ferritin. Recent advances in iron supplementation and nutrient encapsulation and delivery are highlighted. Finally, the current challenges and future developments for multifunctional applications of ferritins are discussed. Expected final online publication date for the Annual Review of Food Science and Technology, Volume 14 is March 2023. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

Effects of heat treatment on the structure, digestive property, and absorptivity of holoferritin

Ferritin, as an iron storage protein, has been considered to be a well-utilized iron supplement. However, the typical thermal processing of food rich in ferritin may affect the structure and function of ferritin as an iron supplement. Here, a plant ferritin (soybean seed ferritin, SSF) and an animal ferritin (donkey spleen ferritin, DSF) were used to analyze the changes in fundamental structure and iron content after thermal treatments (68 °C for 30 min, 100 °C for 10 min). Then, SSF and DSF after thermal treatment were administered intragastrically to mice to further evaluate its digestive stability and absorptivity after thermal processing. Results showed the secondary structure, oligomeric states, iron content, and digestive stability of DSF were maintained better than that of SSF after thermal treatments, indicating that DSF has a higher thermostability than SSF. Both SSF and DSF after thermal treatment exhibited higher absorptivity than untreated ferritins. SSF showed higher absorptivity than DSF whether heated or not.

Extension Peptide of Plant Ferritin from Setaria italica Presents a Novel Fold

The extension peptide (EP) is the most distinctive feature of mature plant ferritin. Some EPs have exhibited serine-like protease activity, which is associated with iron uptake and release. EP forms a helix and a long loop, followed by a conserved core helical bundle. However, whether the EP adopts a stable or uniform folding pattern in all plants remains unclear. To clarify this, we investigated the crystal structure of ferritin-1 from Setaria italica (SiFer1), a type of monocotyledon. In our structure of SiFer1, the EP is different from other EPs in other solved structures of plant ferritins and consisted of a pair of β-sheets, a shorter helix, and two loops, which masks two hydrophobic pockets on the outer surface of every subunit. Furthermore, sequence analysis and structure comparison suggest that the EPs in ferritins from monocotyledons may adopt a novel fold pattern, and the conformations of EPs in ferritins are alterable among different plant species. Furthermore, additional eight iron atoms were first founded binding in the fourfold channels, demonstrating the vital function of fourfold channels in iron diffusion. In all, our structure provides new clues for understanding plant ferritins and the functions of the EP.

Succinylated ferritin as a novel nanocage-like vehicle of polyphenol: Structure, stability, and absorption analysis

Ferritin, a protein with an 8-nm cage structure, can encapsulate and deliver bioactive molecules. In this study, succinylation was adopted to modify plant ferritin to fabricate succinylated red been ferritin (SRBF) at pH 8.0. The SRBF was retained as a cage-like shape (12 nm diameter), while its secondary structure was altered, rendering higher negative charge accompanies by decreased surface hydrophobicity. The SRBF also demonstrated favorable property of reversible assembly regulated by pH-transitions (pH 2.0/7.0), thus enabled successful encapsulation of epigallocatechin gallate (EGCG) for fabrication of EGCG-loaded SRBF complexes with a diameter of ~12 nm. Succinylation enhanced the thermal stabilities of ferritin and the embedded EGCG. Moreover, SRBF markedly improved the transport efficiency of EGCG in Caco-2 monolayers relative to EGCG and that encapsulated in unmodified ferritin. These findings have extended the succinylation reaction for the cage-like protein modification, and facilitated the usage of ferritin variant in delivery of bioactive molecules.

Thermal Treatment Greatly Improves Storage Stability and Monodispersity of Pea Seed Ferritin.

Plant ferritin in holo form is considered as a novel, ideal iron supplement for human nutrition in the 21st century, but its self-degradation and self-association features limit its application on account of the presence of extension peptide (EP), a specific domain only found in plant ferritin. Although reported chemicals such as Phenylmethanesulfonyl fluoride (PMSF) can inhibit its self-degradation, they are not edible and toxic. In the present work, we found that thermal treatment of pea seed ferritin (PSF) in the range of 60 to 80 °C can prolong the storage time of PSF from 3 days to at least 10 days. In the meanwhile, the aggregated form can be inhibited upon such treatment, therefore promoting its monodispersity. More important, such treatment had little effect on the natural shell-like structure of holo PSF and its iron content. In contrast, thermal treatment at higher temperature (90 °C or above) resulted in a change in ferritin structure. These new findings pave the way to the application of plant ferritin as an iron supplement. PRACTICAL APPLICATION: Thermal treatment at 60 to 80 °C can prolong the storage stability of PSF from 3 days to at least 10 days and prevent it from self-aggregation without affecting the shell-like structure. It has been known that the stability of PSF is closely associated with the bioavailability of iron within PSF. From the standpoint of nutrition, the above-mentioned thermal treatment could be used as a cooking method in our daily life or in food industry to improve the bioavailability of ferritin iron, thereby being beneficial for exploration of plant ferritin as a novel, ideal iron supplement to fight against IDA.

Possible role of iron containing proteins in physiological responses of soybean to static magnetic field

Iron is a component of many proteins that have crucial roles in plant growth and development, such as ferritin and catalase. Iron also, as a ferromagnetic element, is assumed to be influenced by a static magnetic field (SMF). In the present study, we examined the relationship between ferrous content and gene expression and activity of ferritin and catalase in soybean plants under the influence of 0, 20, and 30 mT SMF for 5 day, 5 h each. Exposure to 20 mT decreased gene expression of Fe transporter, ferrous and H2O2 contents and gene expression, content and activity of ferritin and catalase. Opposite responses were observed under 30 mT treatments. The results suggest that SMF triggered a signaling pathway that is mediated by iron. The structure and activity of purified ferritin and apoferritin from horse spleen, and catalase from bovine liver proteins under SMF were evaluated as well. Secondary structure of proteins were not influenced by SMF (evidenced by far-UV circular dichroism), whereas their tertiary structure, size, and activity were altered (shown by fluorescence spectroscopy and dynamic light-scattering). From these results, it is likely that the number of iron atoms is involved in the nature of influence of SMF on protein structure.

Iron Release from Soybean Seed Ferritin Induced by Cinnamic Acid Derivatives.

Plant ferritin represents a novel class of iron supplement, which widely co-exists with phenolic acids in a plant diet. However, there are few reports on the effect of these phenolic acids on function of ferritin. In this study, we demonstrated that cinnamic acid derivatives, as widely occurring phenolic acids, can induce iron release from holo soybean seed ferritin (SSF) in a structure-dependent manner. The ability of the iron release from SSF by five cinnamic acids follows the sequence of Cinnamic acid > Chlorogenic acid > Ferulic acid > p-Coumaric acid > Trans-Cinnamic acid. Fluorescence titration in conjunction with dialysis results showed that all of these five compounds have a similar, weak ability to bind with protein, suggesting that their protein-binding ability is not related to their iron release activity. In contrast, both Fe2+-chelating activity and reducibility of these cinnamic acid derivatives are in good agreement with their ability to induce iron release from ferritin. These studies indicate that cinnamic acid and its derivatives could have a negative effect on iron stability of holo soybean seed ferritin in diet, and the Fe2+-chelating activity and reducibility of cinnamic acid and its derivatives have strong relations to the iron release of soybean seed ferritin.

Structural and functional characterization of legume seed ferritin concentrates

The aim of this work was to produce legume seed ferritin concentrates, which were then evaluated for structural properties and susceptibility to gastrointestinal proteases. Five legume seeds, namely red lentil (RL), chickpea (CP), green lentil whole (GLW), moon dal washed (MDW), and yellow split peas (YSP) were subjected to aqueous extraction followed by MgCl2 + sodium citrate-induced precipitation of iron-binding proteins. Ferritin concentrate gross yield was highest for CP (19.1 ± 2.6%) followed by RL (17.6 ± 1.4%), YSP (14.8 ± 1.4%), and GLW (12.9 ± 4.3%), while MDW (9.7 ± 3.3%) was lowest. Iron content of the ferritins ranged from 30–45 mg/100 g in comparison to 5–17 mg/100 g in the seed flour. During simulated gastrointestinal digestion, most of the iron was released by pepsin digestion. We conclude that these iron-enriched protein concentrates are potential ingredients to formulate functional foods and nutraceuticals against iron deficiency anemia. Practical applications Iron deficiency anemia (IDA) continues to be a global problem that can inflict severe damages to human health. The current and most common therapeutic approach to IDA treatment involves the use of inorganic iron supplements. However, the low level of inorganic iron absorption creates serious side-effects such as constipation and high oxidative stress that could lead to degenerative diseases. Organic iron in the form of plant ferritins provides a more absorbable alternative to inorganic iron. However, phytoferritins are present in very small quantities, which make the use of pure forms mostly impracticable. In this work, the preparation of crude phytoferritin concentrates was achieved in addition to their structural and functional characterization that demonstrates potential utility in IDA treatment. The crude ferritin concentrates can be used to formulate functional foods and nutraceuticals as therapies for IDA management.

Directed plant cell-wall accumulation of iron: embedding co-catalyst for efficient biomass conversion.

BackgroundPlant lignocellulosic biomass is an abundant, renewable feedstock for the production of biobased fuels and chemicals. Previously, we showed that iron can act as a co-catalyst to improve the deconstruction of lignocellulosic biomass. However, directly adding iron catalysts into biomass prior to pretreatment is diffusion limited, and increases the cost of biorefinery operations. Recently, we developed a new strategy for expressing iron-storage protein ferritin intracellularly to accumulate iron as a catalyst for the downstream deconstruction of lignocellulosic biomass. In this study, we extend this approach by fusing the heterologous ferritin gene with a signal peptide for secretion into Arabidopsis cell walls (referred to here as FerEX).ResultsThe transgenic Arabidopsis plants. FerEX. accumulated iron under both normal and iron-fertilized growth conditions; under the latter (iron-fertilized) condition, FerEX transgenic plants showed an increase in plant height and dry weight by 12 and 18 %, respectively, compared with the empty vector control plants. The SDS- and native-PAGE separation of cell-wall protein extracts followed by Western blot analyses confirmed the extracellular expression of ferritin in FerEX plants. Meanwhile, Perls' Prussian blue staining and X-ray fluorescence microscopy (XFM) maps revealed iron depositions in both the secondary and compound middle lamellae cell-wall layers, as well as in some of the corner compound middle lamella in FerEX. Remarkably, their harvested biomasses showed enhanced pretreatability and digestibility, releasing, respectively, 21 % more glucose and 34 % more xylose than the empty vector control plants. These values are significantly higher than those of our recently obtained ferritin intracellularly expressed plants.ConclusionsThis study demonstrated that extracellular expression of ferritin in Arabidopsis can produce plants with increased growth and iron accumulation, and reduced thermal and enzymatic recalcitrance. The results are attributed to the intimate colocation of the iron co-catalyst and the cellulose and hemicellulose within the plant cell-wall region, supporting the genetic modification strategy for incorporating conversion catalysts into energy crops prior to harvesting or processing at the biorefinery.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0639-2) contains supplementary material, which is available to authorized users.

Effect of the structure of gallic acid and its derivatives on their interaction with plant ferritin

Gallic acid and its derivatives co-exist with protein components in foodstuffs, but there is few report on their interaction with proteins. On the other hand, plant ferritin represents not only a novel class of iron supplement, but also a new nanocarrier for encapsulation of bioactive nutrients. However, plant ferritin is easy to be degraded by pepsin in the stomach, thereby limiting its application. Herein, we investigated the interaction of gallic acid and its derivatives with recombinant soybean seed H-2 ferritin (rH-2). We found that these phenolic acids interacted with rH-2 in a structure-dependent manner; namely, gallic acid (GA), methyl gallate (MEGA) and propyl gallate (PG) having three HO groups can bind to rH-2, while their analogues with two HO groups cannot. Consequently, such binding largely inhibited ferritin degradation by pepsin. These findings advance our understanding of the relationship between the structure and function of phenolic acids.

Soybean Ferritin Expression in Saccharomyces cerevisiae Modulates Iron Accumulation and Resistance to Elevated Iron Concentrations.

Fungi, including the yeast Saccharomyces cerevisiae, lack ferritin and use vacuoles as iron storage organelles. This work explored how plant ferritin expression influenced baker's yeast iron metabolism. Soybean seed ferritin H1 (SFerH1) and SFerH2 genes were cloned and expressed in yeast cells. Both soybean ferritins assembled as multimeric complexes, which bound yeast intracellular iron in vivo and, consequently, induced the activation of the genes expressed during iron scarcity. Soybean ferritin protected yeast cells that lacked the Ccc1 vacuolar iron detoxification transporter from toxic iron levels by reducing cellular oxidation, thus allowing growth at high iron concentrations. Interestingly, when simultaneously expressed in ccc1Δ cells, SFerH1 and SFerH2 assembled as heteropolymers, which further increased iron resistance and reduced the oxidative stress produced by excess iron compared to ferritin homopolymer complexes. Finally, soybean ferritin expression led to increased iron accumulation in both wild-type and ccc1Δ yeast cells at certain environmental iron concentrations. Iron deficiency is a worldwide nutritional disorder to which women and children are especially vulnerable. A common strategy to combat iron deficiency consists of dietary supplementation with inorganic iron salts, whose bioavailability is very low. Iron-enriched yeasts and cereals are alternative strategies to diminish iron deficiency. Animals and plants possess large ferritin complexes that accumulate, detoxify, or buffer excess cellular iron. However, the yeast Saccharomyces cerevisiae lacks ferritin and uses vacuoles as iron storage organelles. Here, we explored how soybean ferritin expression influenced yeast iron metabolism, confirming that yeasts that express soybean seed ferritin could be explored as a novel strategy to increase dietary iron absorption.

Analysis of Arabidopsis thaliana atfer4-1, atfh and atfer4-1/atfh mutants uncovers frataxin and ferritin contributions to leaf ionome homeostasis

Ferritins are iron-storage proteins involved in the environmental and developmental control of the free iron pool within cells. Plant ferritins are targeted to mitochondria as well as to chloroplasts. AtFer4 is the Arabidopsis thaliana ferritin isoform that can be also targeted to mitochondria. Frataxin is a mitochondrial protein whose role is essential for plants; lack of AtFH frataxin causes early embryo-lethality in Arabidopsis. Because of that, the Arabidopsis atfh KO mutant is propagated in heterozygosis.For exploring the functional interaction between frataxin and ferritin, Arabidopsis double mutant atfer4-1/atfh was isolated and its physiological parameters were measured, as well as its ionome profile, together with those of both atfer4 and atfh single mutants, in different conditions of Fe supply.Impairment of both ferritin and frataxin did not lead to any effect on mitochondrial respiration. However, ionomics revealed that the content of macro- and microelements, occurring when the nutritional Fe supply changes, were altered in the mutants analysed. These results suggest that both ferritin and frataxin can contribute to the composition of the leaf ionome and also confirm ionomics as an excellent tool for detecting alterations in the plant's physiology.

Bioavailability of iron from plant and animal ferritins

Iron deficiency is a major public health problem in the world. Ferritin is being explored as a novel and natural strategy for iron supplementation. The objective of this study was to evaluate iron bioavailability from ferritin isolated from plant and animal sources. The stability of plant ferritin and animal ferritin and the effects of cooking and skim milk were studied by in vitro and in vivo digestion. Results from SDS‐PAGE and Western blot indicate that both plant ferritin and animal ferritin can resist digestion (both under acidic and moderately acidic conditions). Furthermore, ferritin was labelled with 59Fe and bioavailability of iron from ferritin was assessed by uptake into Caco‐2 cells. Our results indicate that iron is taken up from the ferritins and that iron bioavailability from soybean ferritin (rH‐1: rH‐2= 1:1) is the highest. These results may be explained by the binding of ferritin to Caco‐2 cells, which can be attributed to the interaction between ferritin and its putative receptor(s) at the surface of Caco‐2 cells. In conclusion, ferritin from plant and animal sources may be developed as an iron source.

Plant ferritin--a source of iron to prevent its deficiency.

Iron deficiency anemia affects a significant part of the human population. Due to the unique properties of plant ferritin, food enrichment with ferritin iron seems to be a promising strategy to prevent this malnutrition problem. This protein captures huge amounts of iron ions inside the apoferritin shell and isolates them from the environment. Thus, this iron form does not induce oxidative change in food and reduces the risk of gastric problems in consumers. Bioavailability of ferritin in human and animal studies is high and the mechanism of absorption via endocytosis has been confirmed in cultured cells. Legume seeds are a traditional source of plant ferritin. However, even if the percentage of ferritin iron in these seeds is high, its concentration is not sufficient for food fortification. Thus, edible plants have been biofortified in iron for many years. Plants overexpressing ferritin may find applications in the development of bioactive food. A crucial achievement would be to develop technologies warranting stability of ferritin in food and the digestive tract.

Bioavailability of iron from plant and animal ferritins

Iron deficiency is a major public health problem in the world. Ferritin is being explored as a novel and natural strategy for iron supplementation. The objective of this study was to evaluate iron bioavailability from ferritin isolated from plant and animal sources. The stability of plant ferritin and animal ferritin was studied by in vitro and in vivo digestion to determine whether these ferritins can pass through the gastrointestinal tract in intact form. Results from sodium dodecyl sulfate polyacrylamide gel electrophoresis and Western blot indicate that both plant ferritin and animal ferritin can resist digestion (both under acidic and moderately acidic conditions). Furthermore, ferritin was labeled with 59Fe, and bioavailability of iron from ferritin was assessed by uptake into Caco-2 cells. Our results indicate that iron is taken up from the ferritins and that iron bioavailability from soybean ferritin (rH-1:rH-2=1:1) is the highest. These results may be explained by the binding of ferritin to Caco-2 cells, which can be attributed to the interaction between ferritin and its putative receptor(s) at the surface of Caco-2 cells. In conclusion, ferritin from plant and animal sources may be developed as an iron source.

A novel homopolymeric phytoferritin from chickpea seeds with high stability

Iron deficiency is a major public health problem in the world. Phytoferritin as an alternative iron supplement has received increasing attentions recently. Plant ferritins are usually heteropolymers comprising two different H-type subunits, H-1 and H-2, while homopolymeric plant ferritin is rare. In the present study, a newly homopolymeric ferritin chickpea seed ferritin (CSF) was isolated and purified to homogeneity from chickpea seed (Cicer arietinum L.) by two consecutive anion exchange and size exclusion chromatography. SDS-PAGE result indicates that CSF only consists of 28.0 kDa (H-2) subunits, which was identified by Western blot analysis as well. N-terminal sequence and MALDITOF-MS analyses indicate that the subunit of CSF and H-2 subunit of heteropolymeric pea seed ferritin (PSF) share high identity in amino acid sequence. Subsequently, we demonstrated that homopolymeric CSF exhibits a higher catalyzing activity than heteropolymeric PSF at both low and high iron loadings. More importantly, the stability of homopolymeric CSF is much higher than all known plant ferritin due to its highest H-2 subunit content, which is favorable for its application as iron supplement.

Effect of iron status in rats on the absorption of metal ions from plant ferritin.

An isolate of lead-ferritin obtained from soybean seeds sprouted in 25 mM of PbNO3 was introduced into the diet of both iron-deficient and iron non-deficient male rats. After a 21-day administration period, statistical differences in the lead accumulation in the femurs of the rats were noted. Iron-deficient rats accumulated more than four times the amount of lead in their bones than rats without iron-deficiency. No further decrease was observed in haemoglobin concentrations in the groups of animals fed with lead isolates, either iron-deficient or iron non-deficient. Also, no differences in the mean corpuscular haemoglobin (MCH) and mean corpuscular volume (MCV) were observed at the end of the experiment in the group of iron non-deficient rats fed with lead-ferritin isolate compared to the control group of iron non-deficient rats. In the iron-deficient group fed with lead-ferritin isolate, a small increase in haemoglobin concentrations, MCH, MCV and mean corpuscular haemoglobin concentrations (MCHC) was recorded. The results presented in this paper confirm that lead from the tested preparation—lead ferritin isolate—was better absorbed by those rats with induced iron deficiency anaemia. Additionally, we may also suspect based on the obtained results that absorption of ferritin-iron depends on iron status in the body.Electronic supplementary materialThe online version of this article (doi:10.1007/s11130-014-0413-1) contains supplementary material, which is available to authorized users.

Four-Fold Channels Are Involved in Iron Diffusion into the Inner Cavity of Plant Ferritin

From an evolutionary point of view, plant and animal ferritins arose from a common ancestor, but plant ferritin exhibits different features as compared with the animal analogue. One major difference is that the 4-fold channels naturally occurring in plant ferritin are hydrophilic, whereas the 4-fold channels in animal ferritin are hydrophobic. Prior to this study, however, the function of the 4-fold channels in oxidative deposition of iron in phytoferritin remained unknown. To elucidate the role of the 4-fold channels in iron oxidative deposition in ferritin, three mutants of recombinant soybean seed H-2 ferritin (rH-2) were prepared by site-directed mutagenesis, which contained H193A/H197A, a 4-fold channel mutant, E165I/E167A/E171A, a 3-fold channel mutant, and E165I/E167A/E171A/H193A/H197A, where both 3- and 4-channels were mutated. Stopped-flow, electrode oximetry, and transmission electron microscopy (TEM) results showed that H193A/H197A and E165I/E167A/E171A exhibited a similar catalyzing activity of iron oxidation with each other, but a pronounced low activity compared to rH-2, demonstrating that both the 4-fold and 3-fold hydrophilic channels are necessary for iron diffusion in ferritin, followed by oxidation. Indeed, among all tested ferritin, the catalyzing activity of E165I/E167A/E171A/H193A/H197A was weakest because its 3- and 4- fold channels were blocked. These findings advance our understanding of the function of 4-fold channels of plant ferritin and the relationship of the structure and function of ferritin.