18 : 1/ Enzymes: Kinetics of Oxidations 



335 



presence of an excess of hydrogen peroxide, an entirely different absorp- 

 tion spectrum is developed. In contrast to the first complex, the second 

 one appears red. It is enzymatically inactive. Both types of complexes 

 are easier to study with either methyl hydrogen peroxide, CH 3 OOH, or 

 ethyl hydrogen peroxide, C 2 H 5 OOH. Catalase reacts with these two 

 alkyl peroxides to form both the green and the red complexes, but it 

 does not decompose the alkyl peroxides. Spectra for the complexes with 

 methyl hydrogen peroxide are shown in Figure 3. 



Catalase reactions are convenient to study spectrophotometrically for 

 another reason. Besides the possibility for rapid observations afforded 



^ 



<§• 



Reduced 

 Oxidized 



200 



600 



Figure 2. Spectra of the oxidized and reduced forms of an 

 algal cytochrome, Porphyra tenera, cytochrome 553. Spectra 

 from solutions of crystalized enzyme. After S. Katoh, 

 "Crystallization of an Algal Cytochrome," Nature 186: 138 

 (1960). 



by the spectral absorption changes of the enzyme, the peroxide concen- 

 trations can also be observed by measuring the absorption in the ultra- 

 violet region. The spectra of peroxides do not show sharp bands, but 

 rather a curve which rises steadily as the wavelength is decreased from 

 300 m^ to less than 200 m/x. Because many proteins have a minimum 

 in their absorption around 250-230 m/x, this wavelength band has been 

 used to measure the peroxide concentration. Finally, in the peroxidatic 

 reactions, it is often possible to observe the oxidation of AH 2 to A in 

 terms of spectrophotometry changes. 



Such studies have shown that the peroxidatic reaction can be repre- 

 sented by the stoichiometric equations 



e — p x kx p 



E + S^E-S 



fca 



p a k, 



E-S + AH 2 - 



E + A + Products 



