DNA Damage by 5-Aminolevulinic and 4,5-Dioxovaleric Acids in the Presence of Ferritin

Abstract

Cellular accumulation of 5-aminolevulinic acid (ALA), the first specific intermediate of heme biosynthesis, is correlated in liver biopsy samples of acute intermittent porphyria affected patients with an increase in the occurrence of hepatic cancers and the formation of ferritin deposits in hepatocytes. 5-Aminolevulinic acid is able to undergo enolization and to be subsequently oxidized in a reaction catalyzed by iron complexes yielding 4,5-dioxovaleric acid (DOVA). The released superoxide radical (O•−2) is involved in the formation of reactive hydroxyl radical (•OH) or related species arising from a Fenton-type reaction mediated by FeII and CuI. This leads to DNA oxidation. The metal catalyzed oxidation of ALA may be exalted by the O•−2 and enoyl radical-mediated release of FeII ions from ferritin. We report here the potentiating effect of ferritin on the ALA-mediated cleavage of plasmid DNA and the enhancement of the formation of 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodGuo). Plasmid pBR322 was incubated with ALA and varying amounts of purified ferritin. DNA damage was assessed by gel electrophoresis analysis of the open and the linear forms of the plasmid from the native supercoiled structure. Addition of either the DNA compacting polyamine spermidine or the metal chelator ethylenediaminetetraacetic acid (EDTA) inhibited the damage. It was also shown that ALA in the presence of ferritin is able to increase the oxidation of the guanine moiety of monomeric 2′-deoxyguanosine (dGuo) and calf thymus DNA (CTDNA) to form 8-oxodGuo as inferred from high performance liquid chromatography (HPLC) measurements using electrochemical detection. The formation of the adduct dGuo-DOVA wasdetected in CTDNA upon incubation with ALA and ferritin. In a subsequent investigation, the aldehyde DOVA was also able to induces strand breaks in pBR322 DNA.