SOLUBILIZATION AND PROPERTIES OF THE DPNH DEHYDROGENASE III 



at 30°, pH 7-4 (Fig. 8). It should be noted that this DPNH concentration 

 is only about one-half of that which gives apparent optimal activity (Fig. 5) 

 in the ferricyanide assay. While an increase in the initial DPNH concentra- 



Effect of DPN 



10- 



DPNH =1 5xI0"''m 

 + 9xI0"'m DPN 



No DPN 



2 4 6 8 10 



l/m"L. Fe(CN)^"" 



Fig. 7. Competitive inhibition of soluble, purified dehydrogenase by 9 x 10" 

 M DPN. Standard ferricyanide assay; DPXH concentration = i -5 x 10"* M. 



M DPNH X 10^ 



Fig. 8. Lineweaver-Burk plot of effect of DPNH concentration on activity. 

 The values on the ordinate are maximal velocities of ferricyanide reduction 

 (^max with respect to ferricvanide) corresponding to each concentration of 

 DPNH at pH 74, 30 . 



tion beyond about 2 x lo-"* m fails to increase the measured rate, even 

 under conditions where the inhibitory effect of excess substrate is elimina- 

 ted, it is doubtful if this value represents a true "saturation" of the dehy- 

 drogenase. 



