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1342 BIOCHEMICAL SOCIETY TRANSACTIONS Model Compounds with Superoxide Dismutase Activity : Iron Porphyrins and other Iron Complexes BARRY HALLIWELL and ROBERT F. PASTERNACK Department of Biochemistry, University of London King's College, Strand, London WC2R 2LS, U.K. The enzyme superoxide dismutase (EC 1.15.1.1) is essential to aerobic life (Fridovich, 1975; Halliwell, 1978). All superoxide dismutases are metalloproteins, containing manganese, iron or copper and zinc as the prosthetic groups. However, some simple metal complexes also react with the superoxide radical (02-'); for example, complexes of Cu(I1) with amino acids (Brigelius et al., 1974) and complexes of Mn(1I) with pyrophosphate (Kono etal., 1976) or with quinolino compounds (Howie et al., 1977). Complexes of Cu(I1) 0 1 Mn(I1) with EDTA do not react with 02-.(Halliwell, 1975). In contrast, complexes of Fe(I1) or Fe(II1) with EDTA react rapidly with 02-* (Halliwell, 1975). By competition of Fe(I1)-EDTA with Nitro Blue Tetrazolium for 02-* generated in a xanthine/xanthine oxidase system at pH 10.1, a rate constant of about 3 x 105~-'*s-' was calculated for the reaction of Fe(l1)-EDTA with 02-'at 20°C. Complexes of iron(I1) or iron(II1) with diethylenetriaminepenta-acetic acid also react with 02-.at pH10.1 (k 1 x 105~-'.s-' at 20°C). Halliwell(l975) reported that haematin reacts with 02-* at pH 10, but under the conditions used this compound would have been extensively aggregated. 0 2 -is. known to reduce iron(II1) protoporphyrin IX dimethyl ester in organic solvents (Hill et al., 1974). Certain protein-porphyrin complexes interact with 02-* in aqueous solution, including oxidized cytochrome c ( k 1.1 x 106~vl'.s-'at pH7.2: Koppenol et a/., 1976), horseradish peroxidase (k 1 x 10'-1 x 1 0 9 ~ - ' . s - ' at pH5; Sawada & Yamazaki, 1973), oxyhaemoglobin and methaemoglobin ( k - 4 x lo3 and 6x 103~-1.s-L at pH7; Sutton et al., 1976). However, catalase does not react with 02-'at either pH7.8 or 10.2 (Halliwell, 1973). As part of an investigation into the relation between the structure of porphyrins and their reaction with 02-., we have studied the properties of various metal-ion complexes of the water-soluble porphyrin tetrakis-(4-N-methyl)pyridylporphine (Fig. 1). The spectral and aggregation properties of this compound are well-known (Pasternack eta/., 1977). The compound itself was found not to react with 02-'in a xanthine/xanthine oxidase system at pH1O.l, nor did its complexes with Zn(1I) or Cu(l1). The Co(ll1) derivative of tetrakis-(4-N-methyl)pyridylporphinereacted slowly with O2-., but it could not be decided if the reaction was catalytic in our system. However, the iron(lI1) - -- - Fig. 1. Structure of tetrakis-(4-N-methyl)pyridyIporphine iron(IZZ) 1978 577th MEETING, OXFORD 1343 complex reacted rapidly with 02-.in a catalytic manner. The rate constant for O,-* dismutation was determined by competition of the iron porphyrin with Nitro Blue Tetrazolium in a xanthine/xanthine oxidase system at pH 10.1 ;a value of 3 x ~O'M-' .s-' at 20°C was obtained. The iron porphyrin had no effect on xanthine oxidase activity. Under our reaction conditions, at least 14 molecules of Oz-' reacted with each molecule of iron porphyrin. Unless catalase was added to the reaction mixture, the H2O2generated caused degradation of the iron porphyrin to a product with a much lower Soret intensity, probably a bile-pigment type of compound. This product also catalysed dismutation of 02-' ( k = 2 X 1o6M-' .S-' ). In the presence of catalase, the Soret band of the iron porphyrin decreased and broadened slightly on first exposure to xanthine/xanthine oxidase, but then re-attained its original intensity. This suggests the intermediate formation of some iron(I1) porphyrin. The simplest way of representing the reaction of tetrakis-(4-N-methyl)pyridylporphineiron( 111) [Fe(III)TMpyP] with 0,-. would be: Fe(II1)TMpyP +02-' --> Fe(II)TMpyP+O, Fe(II)TMpyP+O2-'+2HZ0 Fe(III)TMpyP+H20,+20H- (1) (2) Further investigations on the reaction mechanism and on the effect of complexformation of the porphyrin with proteins are required. Brigelius, R., Spottl, R., Bors, W., Lengfelder, E., Saran, M. & Weser, U. (1974) FEBS Left. 47, 72-75 Fridovich, 1. (1975) Annu. Rev. Biochem. 44, 147-159 Halliwell, B. (1973) Biochetn. 1.135, 379-381 Halliwell, B. (1975) FEBS Lrft. 56, 34-38 Halliwell, B. (1978) Cell B i d . Int. Rep. 2 , 113-128 Hill, H. A. O . ,Turner, D. R. & Pellizer, G. (1974) Biochem. Biophys. Res. Commun. 56,739-744 Howie, J. K., Morrison, M. K. & Sawyer, D. T. (1977) in E/ecfrochemica/SfuciesofBio/ogica/ Sysferns (Sawyer, D. T., ed.), pp. 97-1 12, American Chemical Society, Washington Kono, Y., Takahashi, M. & Asada, K. (1976) Arch. Biochem. Biophps. 174, 454-462 Koppenol, W. H., Van Buuren, K . J. 449, 157-168 Pasternack, R. F., Lee, H., Malek, P. Sawada, Y . & Yamazaki, 1. (1973) Biochim. Biophys. Acfa 327, 257-265 Sutton, H. C., Roberts, P. B. & Winterbourn, C. C. (1976) Biochem. J . 155, Oxidation of Thiol Compounds by Catalase and Peroxidase in the Presence of Manganese@) and Phenols J O H A N D E RYCKER and BARRY HA.LL1WELL Department of Biochemistry, University of London King's College, Strand, London WC2R 2LS, U.K. Catalase breaks down H 2 0 2to O2 and water, but in the presence of H 2 0 2 it can also act as a peroxidase, oxidizing compounds such as ethanol, formate and nitrite. O n exposure to acid o r alkaline p H values, the catalatic activity of catalase is decreased, but the peroxidatic activity is increased (Inada et al., 1961 ; Marklund, 1973). For example, catalase gains the ability to oxidise NADH in the presence of Mn2+and phenols (Caravca & May, 1964). It also catalyses conversion of 3-hydroxyanthranilate into cinnabarinate in the presence of MnZ+(Savage & Prinz, 1977). Breakdown of H 2 0 2 by catalase is inhibited by thiol compounds (Dale & Russell, 1956; Keilin & Nicholls, 1958; Olinescu & Cotae, 1973; Scheifer et af., 1976). We report here that catalase can bring about oxidation of thiol compounds in the presence of MnZ+ and certain phenols. Horseradish peroxidase catalyses a similar reaction (Stonier & Yang, 1973), but its specificity for phenols is different from that of catalase. Vol. 6