ENZYME-SUBSTRATE COMPOUNDS 



its formation and the latter does not participate in the de- 

 composition of hydrogen peroxide (10). The lifetime of such 

 a ternary complex would be so short (2 X 10~^ sec.) that con- 

 siderations of whether or not the two oxidizing and reducing 

 equivalents of the two peroxide molecules were transferred 

 stepwise or simultaneously begin to lose their importance. But 

 direct spectroscopic studies of strong catalase in several molar 

 peroxide are now needed to determine whether the ternary 

 complex actually exists or whether the reaction is inhibited by 

 the formation of inactive complex II, as has been observed 

 spectroscopically in more dilute peroxide (10). 



PEROXIDASE AND HYDROGEN PEROXmE 



Complexes I of catalase and peroxidase appear to be 

 physically and chemically identical, but their protein parts 

 give rise to different mechanisms and specificities in their 

 reactions with hydrogen donors. It is probable that our data 

 on the nature of the combination of catalase and hydrogen 

 peroxide to form complex I apply equally well to peroxidase 

 and direct experiments over a more limited range support 

 this idea. 



Direct spectroscopic measurement of the combination of 

 peroxidase and hydrogen peroxide carried out in the rapid-flow 

 apparatus has shown that a second-order reaction occurs 

 (ki = 0.9 X 10^ M-^ sec.-i at 25 ° C), but the results have been 

 limited to times longer than 2 millisec. (7). There are in ad- 

 dition a number of assay methods for the over-all activity of 

 peroxidase from which the value of ^i could be computed. 

 The guaiacol assay method for peroxidase fulfills the conditions 

 for determining ^i from the rate of the over-all reaction: 



dx/dt = -T— ^— r; (5) 



— + — 

 kix kiU 



if A:4<3 ^ kxX 



dx/dt = kixe. (6) 



315 



