Understanding Differences in Mechanisms of Action of Fe vs Mn Porphyrins: Comparison of Their Reactivities Towards Cellular Reductants and Reactive Species

Abstract

Understanding Differences in Mechanisms of Action of Fe vs Mn Porphyrins: Comparison of Their Reactivities Towards Cellular Reductants and Reactive Species Artak Tovmasyan, Tin Weitner, Emily Roberts, Melba Jaramillo, Ivan Spasojevic, Kam Leong, Margaret Tome, Ludmil Benov, and Ines Batinic-Haberle Duke University, University of Arizona, Kuwait University, Kuwait Fe and Mn N-substituted pyridylporphyrins (FePs and MnPs) are successfully studied in models of oxidative stress. Replacement of Mn with Fe in MnSOD enzyme results in dramatic change in metal coordination, which shifts M(III)/M(II) reduction potential, E1/2 out of the region optimal for catalysis of O2dismutation, and results in the loss of enzyme activity. In contrast, both FePs and MnPs have nearly identical E1/2 and thus dismute O2 and reduce ONOO with similarly high rate constants. Yet, there is a distinct difference in metal coordination field: at pH 7.8 axial coordination sites of Mn are occupied with two H2O molecules, while Fe bears H2O and OH ligands. The OH ligand neutralizes a positive charge at the Fe site, and labilizes trans-axial H2O, resulting in enhanced reactivity of FePs. FePs allow SOD-deficient E. coli to grow as good as wild type at up to 1000-fold lower concentrations than MnPs, but become toxic at doses where MnPs are efficient. When combined with ascorbate, MnPs become toxic to E. coli, whereas FePs toxicity is reversed. In a different study, both FePs and MnPs exert up to 10-fold higher toxicity towards cancer vs normal cells; however, FePs are cytotoxic at up to 10-fold lower concentrations than MnPs. When given with ascorbate, MnPs are toxic towards cancer cells (and not towards normal cells), while FePs/ascorbate system is toxic to neither cancer nor normal cells. To understand such large difference in bioactivity of FePs vs MnPs, we explored their reactivity towards cellular reductants. Thiol (cysteine and glutathione) oxidase activity, leading to H2O2 production, was revealed for both MnPs and FePs. The FePs are several-fold more reactive than MnPs, resulting in higher production of H2O2. Yet, FePs are very unstable towards H2O2; “free” Fe released by rapid degradation of FePs results in cell death, presumably via Fenton chemistry. In contrast, MnPs produce less H2O2, are more stable towards it, plus “free” Mn does not undergo Fenton chemistry. With ascorbate in cell medium, FePs are rapidly degraded by H2O2 produced in the medium due to FePs/ascorbate cycling, and in turn have small chance to enter and damage the cell. With MnPs/ascorbate, and due to MnPs higher stability, excessive levels of H2O2 are produced, thus accounting for enhanced cytotoxicity. Unlike cancer cells, normal cells are up to 10-fold less sensitive either to porphyrin or its combination with ascorbate due to the abundance of H2O2-removing systems.