Catalase Oxidation Mechanisms 247 



until its electronegativity exceeds that of the ligands, and that the latter 

 provide the next electron to be removed. Especially when a ligand contains a 

 large number of conjugated double bonds, only one or two electrons are 

 likely to be extracted from the metal before the ligand suffers oxidation. 

 Cahill and Taube (1951) were able to show with several metallo-phthalo- 

 cyanines, apparently 4-co-ordinate, that an electron could be removed from 

 the phthalocyanine (£° = —IV approx.) by a 1-electron acceptor. Two- 

 electron oxidants of comparable strength were considerably slower in action. 

 The closely related metallo-porphyrins are expected to behave similarly, 

 and also the 6-co-ordinate forms which occur in catalase, peroxidase, 

 metmyoglobin, etc., but in these latter compounds a higher oxidation 

 state of the metal should be reached before an electron is lost from the 

 porphyrin. 



When metmyoglobin (Mb"^) is oxidized by H.2O2 the solution contains a 

 free radical which has been detected by electron spin resonance absorption 

 (Gibson and Ingram, 1956). In later experiments Gibson, Ingram and 

 Nicholls (1958) showed that the principal oxidation product (Mb^^) is not 

 the free radical, and that the latter is present in about ten times lower concen- 

 tration. We may infer that although when Mb"^ is oxidized by H.^Oo an 

 electron is given up rather more readily by the Fe+++ than by the porphyrin, 

 the latter is the site of attack on some of the Mb^" molecules, either in a side 

 reaction or by electron transfer between myoglobin molecules in different 

 oxidation states. 



It is thus necessary to consider the possibihty that oxidation of catalase 



\ / \ / 

 yields structures of the type -C Fe™, -C Fe^^, etc. (notation of King 



and Winfield, 1959a); the carbon atom shown attached indirectly to Fe 

 represents one of the carbon atoms of a ligand which, before loss of an elec- 



tron, is shown as C Fe"^. Free radical character in the porphyrin is one 



of the possible explanations of the diminished Soret band in Cat. H2O2 I 

 (see, for example, George, 1952). A number of factors count against a free 

 radical structure, however, and these will now be discussed. 



Perhaps the most striking feature of the reactions of catalase and peroxidase 

 is that no one has been able, in the course of numerous oxidations with a 

 variety of agents, to remove one and only one electron from the enzyme 

 molecule. Cat. H2O2 II and the peroxidase-peroxide complex (Per. H0O2 II) 

 have an oxidation state corresponding to one less electron than catalase and 

 peroxidase, but they cannot be formed directly from the latter (Fergusson, 

 1956). Apparently the ligands in catalase and peroxidase are protected by 

 steric hindrance or other means from attack by a 1-electron oxidant, while the 

 iron atom is co-ordinated in such a way that it strongly resists direct electron 



