Oxidative damage to ferritin by 5-aminolevulinic acid.
5-Aminolevulinic acid (ALA), a heme precursor overproduced in various porphyric disorders, has been implicated in iron-mediated oxidative damage to biomolecules and cell structures. From previous observations of ferritin iron release by ALA, we investigated the ability of ALA to cause oxidative damage to ferritin apoprotein. Incubation of horse spleen ferritin (HoSF) with ALA caused alterations in the ferritin circular dichroism spectrum (loss of a alpha-helix content) and altered electrophoretic behavior. Incubation of human liver, spleen, and heart ferritins with ALA substantially decreased antibody recognition (51, 60, and 28% for liver, spleen, and heart, respectively). Incubation of apoferritin with 1-10mM ALA produced dose-dependent decreases in tryptophan fluorescence (11-35% after 5h), and a partial depletion of protein thiols (18% after 24h) despite substantial removal of catalytic iron. The loss of tryptophan fluorescence was inhibited 35% by 50mM mannitol, suggesting participation of hydroxyl radicals. The damage to apoferritin had no effect on ferroxidase activity, but produced a 61% decrease in iron uptake ability. The results suggest a local autocatalytic interaction among ALA, ferritin, and oxygen, catalyzed by endogenous iron and phosphate, that causes site-specific damage to the ferritin protein and impaired iron sequestration. These data together with previous findings that ALA overload causes iron mobilization in brain and liver of rats may help explain organ-specific toxicities and carcinogenicity of ALA in experimental animals and patients with porphyria.
Rocha
Maria E M
ME
Departamento de Bioqui;mica, Instituto de Qui;mica, Universidade de São Paulo, Av. Prof. Lineu Prestes 748, CEP 05508-900, São Paulo, SP, Brazil.
Dutra
Fernando
F
Bandy
Brian
B
Baldini
Regina L
RL
Gomes
Suely L
SL
Faljoni-Alário
Adelaide
A
Liria
Cleber W
CW
Miranda
M Terêsa M
MT
Bechara
Etelvino J H
EJ
D000622:Aminolevulinic Acid / Q000378:metabolism / Q000494:pharmacology*; D000818:Animals; D001052:Apoferritins / Q000737:chemistry / Q000187:drug effects; D002942:Circular Dichroism; D004305:Dose-Response Relationship, Drug; D015180:Electrophoresis, Gel, Two-Dimensional; D005293:Ferritins / Q000737:chemistry / Q000187:drug effects*; D006736:Horses; D006801:Humans; D066298:In Vitro Techniques; D007501:Iron / Q000378:metabolism; D008099:Liver / Q000737:chemistry; D008353:Mannitol / Q000494:pharmacology; D009206:Myocardium / Q000737:chemistry; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D010100:Oxygen / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013154:Spleen / Q000737:chemistry; D013438:Sulfhydryl Compounds / Q000032:analysis; D013481:Superoxides / Q000378:metabolism; D014364:Tryptophan / Q000187:drug effects
D013438:Sulfhydryl Compounds; D013481:Superoxides; D008353:Mannitol; D000622:Aminolevulinic Acid; D014364:Tryptophan; D005293:Ferritins; D001052:Apoferritins; D007501:Iron; D010100:Oxygen