Chem. Pharm. Bull., 48(2),223-230, February 2000

Regular Articles

Mechanism of Superoxide Dismutase-Like Activity of Fe(II) and Fe(III) Complexes of Tetrakis-N, N, N', N'(2-pyridylmethyl)ethylenediamine

Tomohisa HIRANO,a Masaaki HIROBE,a Kazuo KOBAYASHI,b Akira ODANI,c Osamu YAMAUCHI,c Masanori OHSAWA,a Yoshinori SATOW,a and Tetsuo NAGANO*,a

Graduate School of Pharmaceutical Sciences, The University of Tokyo,a 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan, Institute of Scientific and Industrial Research, Osaka University,b 8-1 Mihogaoka, Ibaraki, Osaka 567-0047, Japan, and Department of Chemistry, Faculty of Science,c Nagoya University, Chikusa-ku, Nagoya 464-0814, Japan.

The superoxide dismutase (SOD) activity of iron(II) tetrakis-N, N, N', N'(2-pyridylmethyl)ethylenediamine complex (Fe-TPEN) was reexamined using a pulse radiolysis method. In our previous study (J. Biol. Chem., 264, 9243-9249 (1989)), we reported that this complex has a potent SOD activity in a cyt. c (cytochrome c)-based system (IC50=0.8 mM) and protects E. coli cells against paraquat toxicity. The present pulse radiolysis experiment revealed that Fe(II)TPEN reacts stoichiometrically with superoxide to form Fe(III)TPEN with a second-order rate constant of 3.9×106M-1s-1 at pH 7.1, but superoxide did not reduce Fe(III)TPEN to Fe(II)TPEN. The reaction of Fe(III)TPEN and superoxide was biphasic. In the fast reaction, an adduct (Fe(III)TPEN-superoxide complex) was formed at the second-order rate constant of 8.5×105M-1s-1 at pH 7.4. In the slow one, the adduct reacted with another molecule of the adduct, regenerating Fe(III)TPEN. In the cyt. c method with catalase, this Fe(III)TPEN-superoxide complex showed cyt. c oxidation activity, which had led to overestimation of its SOD activity. Based on the titration data, the main species of complex in aqueous media at neutral pH was indicated to be Fe(III)TPEN(OH-). A spectral change after the reduction with hydrated electron indicates that the OH- ion coordinates directly to Fe(III) by displacing one of the pyridine rings. The X-ray analysis of [Fe(II)TPEN]SO4 supported this structure. From the above results we propose a novel reaction mechanism of FeTPEN and superoxide which resembles a proton catalyzed dismuting process, involving Fe(III)TPEN-superoxide complex.

Key words superoxide dismutase mimics; Tetrakis-N, N, N', N'(2-pyridylmethyl)ethylenediamine; free radical; pulse radiolysis; X-ray analysis; pH-titration