The release of iron from horse spleen ferritin by reduced flavins

Abstract

Ferritin-Fe(III) was rapidly and quantitatively reduced and liberated as Fe(II) by FMNH(2), FADH(2) and reduced riboflavin. Dithionite also released Fe(II) from ferritin but at less than 1% of the rate with FMNH(2). Cysteine, glutathione and ascorbate gave a similar slower rate and yielded less than 20% of the total iron from ferritin within a few hours. The reduction of ferritin-Fe(III) by the three riboflavin compounds gave complex second-order kinetics with overlapping fast and slow reactions. The fast reaction appeared to be non-specific and may be due to a reduction of Fe(III) of a lower degree of polymerization, equilibrated with ferritin iron. The amount of this Fe(3+) ion initially reduced was small, less than 0.3% of the total iron. Addition of FMN to the ferritin-dithionite system enhanced the reduction; this is due to the reduction of FMN by dithionite to form FMNH(2) which then reduces ferritin-Fe(III). A comparison of the thermodynamic parameters of FMNH(2)-ferritin and dithionite-ferritin complex formation showed that FMNH(2) required a lower activation energy and a negative entropy change, whereas dithionite required 50% more activation energy and showed a positive entropy change in ferritin reduction. The effectiveness of FMNH(2) in ferritin-Fe(III) reduction may be due to a specific binding of the riboflavin moiety to the protein portion of the ferritin molecule.

Mobilisation of iron from ferritin fractions of defined iron content by biological reductants

“…The most effective mobilising agent is cysteine, followed by ascorbic acid; glutathione is a very poor reductant of ferritin iron. This is in agreement with earlier results of Mazur et al [7] and Sirivech et al [3]. Yet at concentrations of 5 mM mobilisation of only 12-I 5% of the iron content is obtained in 24 hr at room temperature.…”

“…It does not seem likely that such reducing agents or complexants play a major role in ferritin mobilisation in vivo. By comparison, with FMNHz reduction of ferritin iron is extremely rapid (R. R. Crichton, M. Wauters and F. Roman unpublished work and [3]) 100% of the iron being reduced by 0.2 mM FMNHz in 10 min [3]. The effect of addition of chelating agents together with reducing agents did not increase the rate of iron release appreciably.…”

“…The iron can be released by reduction, for example with sodium dithionite [l] or with thioglycolate [ 21. In the cell ferritin iron seems likely to be mobilised by a reductive system, which may involve FMNHz [3]. It is of interest to establish the nature of the biological reductant which is involved in iron release from ferritin and to analyse the effect of the iron content of the molecule on the rate of iron release.…”

Reductive mobilisation of ferritin iron

“…100 mM [13]. We can observe a more rapid phase at the beginning followed by a phase of slower mobilisation rate (Fig.…”

“…However, the effectiveness strongly varies with the reagent used. Cysteine and glutathione, for example, give poor mobilisation rates [13], while a faster mobilisation is observed with thioglycollic acid as used for the formation of apoferritin from ferritin [35]. This order of reactivities is not compatible with a predominant dependence of reduction rates on the redox potential since the thiol-disulfide couples involving cysteine and glutathione have lower potentials (Eb = -0.34 V) than thioglycollic acid (Eb = -0.14 V) [36].…”