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Evidence of H- and L-chains have co-operative roles in the iron-uptake mechanism of human ferritin.

ABSTRACT: The ability to incorporate iron in vitro was studied in homopolymers of human ferritin L-chain, human ferritin H-chain and its variants and in homopolymer mixtures. The H-chain variants carried amino acid substitutions in the ferroxidase centre and/or in carboxy residues on the cavity surface. Iron incorporation was examined by gel electrophoresis of the reaction products by staining for iron and protein. It was found that inactivation of the ferroxidase centre combined with the substitution of four carboxy groups on the cavity abolished the ability of H-chain ferritin to incorporate iron. Competition experiments with limited amounts of iron showed that, at neutral pH, L-chain ferritin is more efficient in forming iron cores than the H-chain variants altered at the ferroxidase activity or in the cavity. Competition experiments at pH 5.5 demonstrated that L-chain apoferritin is able to incorporate iron only when in the presence of H-chain variants with ferroxidase activity. The results indicate that L-chain apoferritin has a higher capacity than the H-chain apoferritin to induce iron-core nucleation, whereas H-chain ferritin is superior in promoting Fe(II) oxidation. The finding of cooperative roles of the H- and L-chains in ferritin iron uptake provides a clue to understanding the biological function of isoferritins.

SUBMITTER: Levi S

PROVIDER: S-EPMC1132051 | biostudies-other | 1992 Dec

REPOSITORIES: biostudies-other

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Project description:Background:Iron deficiency is an enduring global health problem that requires new remedial approaches. Iron absorption from soybean-derived ferritin, an ?550-kDa iron storage protein, is comparable to bioavailable ferrous sulfate (FeSO4). However, the absorption of ferritin is reported to involve an endocytic mechanism, independent of divalent metal ion transporter 1 (DMT-1), the transporter for nonheme iron. Objective:Our overall aim was to examine the potential of purified ferritin from peas (Pisum sativum) as a food supplement by measuring its stability under gastric pH treatment and the mechanisms of iron uptake into Caco-2 cells. Methods:Caco-2 cells were treated with native or gastric pH-treated pea ferritin in combination with dietary modulators of nonheme iron uptake, small interfering RNA targeting DMT-1, or chemical inhibitors of endocytosis. Cellular ferritin formation, a surrogate measure of iron uptake, and internalization of pea ferritin with the use of specific antibodies were measured. The production of reactive oxygen species (ROS) in response to equimolar concentrations of native pea ferritin and FeSO4 was also compared. Results:Pea ferritin exposed to gastric pH treatment was degraded, and the released iron was transported into Caco-2 cells by DMT-1. Inhibitors of DMT-1 and nonheme iron absorption reduced iron uptake by 26-40%. Conversely, in the absence of gastric pH treatment, the iron uptake of native pea ferritin was unaffected by inhibitors of nonheme iron absorption, and the protein was observed to be internalized in Caco-2 cells. Chlorpromazine (clathrin-mediated endocytosis inhibitor) reduced the native pea ferritin content within cells by ?30%, which confirmed that the native pea ferritin was transported into cells via a clathrin-mediated endocytic pathway. In addition, 60% less ROS production resulted from native pea ferritin in comparison to FeSO4. Conclusion:With consideration that nonheme dietary inhibitors display no effect on iron uptake and the low oxidative potential relative to FeSO4, intact pea ferritin appears to be a promising iron supplement.

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Evidence of H- and L-chains have co-operative roles in the iron-uptake mechanism of human ferritin.

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