Mechanism of the transition from plant ferritin to phytosiderin.
Soluble and insoluble forms of ferritins have been purified from dry pea seeds by gel filtration. The insoluble form is called phytosiderin by analogy with animal hemosiderin. Native gel electrophoresis of these two forms have shown that the soluble one (ferritin) is homogenous in size and more compact than the insoluble one (phytosiderin) which is heterogenous in size. However, when iron is removed from these two classes of molecules (apoferritin), they have the same mobility in isopycnic centrifugations. Polyacrylamide-sodium dodecyl sulfate gel electrophoresis revealed a difference in their subunit composition: ferritin molecules are built up from a 28-kDa subunit and phytosiderin from a 26.5-kDa subunit. Partial proteolysis using a Staphylococcus aureus protease indicates a strong relationship between these two polypeptides. Intermediates between these two forms have also been characterized and are composed of both subunits in various amounts. Ferritin and phytosiderin are both able to incorporate iron in vitro into their mineral core. It is also shown that in vitro iron exchange induces ferritin degradation. This degradation is prevented by inhibitors of the Fenton cycle (iron chelates like o-phenanthroline and desferrioxamine B) and reduced by Tris, a radical scavenger. Under in vitro conditions of controlled radical damage the 28-kDa subunit is converted into the 26.5-kDa subunit. Purification of the 28-kDa subunit has allowed us to determine the NH2-terminal sequence. The NH2 extremity of the 26.5-kDa subunit is heterogenous, but the sequence of its main component is identical to the sequence of the 28-kDa subunit downstream residue Leu-21. These data indicate that the 26.5-kDa subunit is produced by radical mediated damage leading to a series of cleavages in the NH2 terminal part of the 28-kDa subunit.
Laulhere
J P
JP
Laboratoire de Biologie Moléculaire Végétale, Centre National de la Recherche Scientifique UA 1178, Université Joseph Fourier (Grenoble I), France.
Laboure
A M
AM
Briat
J F
JF
D000595:Amino Acid Sequence; D002499:Centrifugation, Density Gradient; D055598:Chemical Phenomena; D002621:Chemistry; D002850:Chromatography, Gel; D004587:Electrophoresis, Agar Gel; D004591:Electrophoresis, Polyacrylamide Gel; D007887:Fabaceae; D005293:Ferritins / Q000302:isolation & purification / Q000378:metabolism; D006878:Hydroxides / Q000494:pharmacology; D017665:Hydroxyl Radical; D007501:Iron / Q000378:metabolism; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments; D010447:Peptide Hydrolases / Q000378:metabolism; D010946:Plants, Medicinal; D012639:Seeds / Q000032:analysis*; D012995:Solubility
D006878:Hydroxides; D046911:Macromolecular Substances; D010446:Peptide Fragments; C058550:phytosiderin; D017665:Hydroxyl Radical; D005293:Ferritins; D007501:Iron; D010447:Peptide Hydrolases