Catalase Oxidation Mechanisms 255 



to times of about 1 millisec and then by studies of the rate laws for peroxide decompo- 

 sition up to about 10 M peroxide (Ogura, Arch. Biochein. Biophys. 57, 288, 1955). No 

 significant deviations occurred for half lives of such complexes of about 5 x 10^* sec 

 or less. This time is long enough, however, to allow intramolecular rearrangements 

 to occur and no definitive conclusions were reached. 



More recently Schonbaum and I have investigated the types of reactions that could 

 be involved in Eq. (1) and (2) in order to determine whether some difference between 

 the two reactions in their dependence on the concentration of methylhydroperoxide 

 would be expected. If catalase forms a complex of peroxide and the enzyme, the usual 

 form of the equilibrium equation should apply. If, however, instead of forming a 

 complex, the components of peroxide are released as an alcohol, a modified equilibrium 

 equation is required which involves a squared term in the intermediate concentration. 

 We have considered several cases as discussed below. 



The terminology used here is illustrated for a simple equilibrium: 



h 

 E + S^ ES (3) 



{e-p) (x) (p) 



x(e — p) 



This represents Eq. (1) above. 



It is perhaps trivial to indicate that a simple equilibrium followed by an irreversible 

 transformation gives no equilibrium: 



E + S^ESj (5) 



ESi-^ESa (6) 



If the irreversible step is followed by a decomposition of the intermediate to the free 

 enzyme, the system simulates an equilibrium but is actually in a steady state. 



(7) 

 (8) 



(9) 



Such a reaction appears to occur in peroxidase where endogenous donor (AHj) 

 participates in the reaction of Eq. (9) and in catalase where endogenous alcohol 

 participates. This equation is valid when such a donor is in excess, i.e. [AHg] = 

 constant. Both these reactions are negligible in pure enzyme preparations which 

 contain no donor, i.e. [AHj] = 0. 



If the components of peroxide are expended in the initial reaction and a hydrogen 

 donor such as an alcohol is formed, reaction (8) may give rise to a simulated equi- 

 librium: 



E + S^ES' + AH2 (10) 



{e-p) (x) (p') (a) 



This represents Eq. (2) above. Here: 



x(e — p') 



