"Opening" the ferritin pore for iron release by mutation of conserved amino acids at interhelix and loop sites.
Ferritin concentrates, stores, and detoxifies iron in most organisms. The iron is a solid, ferric oxide mineral (< or =4500 Fe) inside the protein shell. Eight pores are formed by subunit trimers of the 24 subunit protein. A role for the protein in controlling reduction and dissolution of the iron mineral was suggested in preliminary experiments [Takagi et al. (1998) J. Biol. Chem. 273, 18685-18688] with a proline/leucine substitution near the pore. Localized pore disorder in frog L134P crystals coincided with enhanced iron exit, triggered by reduction. In this report, nine additional substitutions of conserved amino acids near L134 were studied for effects on iron release. Alterations of a conserved hydrophobic pair, a conserved ion pair, and a loop at the ferritin pores all increased iron exit (3-30-fold). Protein assembly was unchanged, except for a slight decrease in volume (measured by gel filtration); ferroxidase activity was still in the millisecond range, but a small decrease indicates slight alteration of the channel from the pore to the oxidation site. The sensitivity of reductive iron exit rates to changes in conserved residues near the ferritin pores, associated with localized unfolding, suggests that the structure around the ferritin pores is a target for regulated protein unfolding and iron release.
Jin
W
W
CHORI (Children's Hospital Oakland Research Institute), 5700 Martin Luther King Jr. Way, Oakland, California 94609, USA.
Takagi
H
H
Pancorbo
B
B
Theil
E C
EC
D020816:Amino Acid Motifs / Q000235:genetics; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D001120:Arginine / Q000235:genetics; D001224:Aspartic Acid / Q000235:genetics; D017124:Conserved Sequence / Q000235:genetics*; D005293:Ferritins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D007501:Iron / Q000378:metabolism*; D007502:Iron Chelating Agents / Q000378:metabolism; D007930:Leucine / Q000235:genetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017510:Protein Folding; D017433:Protein Structure, Secondary / Q000235:genetics; D011892:Rana catesbeiana
D007502:Iron Chelating Agents; D010446:Peptide Fragments; D001224:Aspartic Acid; D005293:Ferritins; D001120:Arginine; D007501:Iron; D007930:Leucine