HUGO THEORELL 



general phenomenon, since it does not occur in yeast ADH, 

 triosephosphate, or glutamic dehydrogenases. 



The displacement of the band enabled us to establish spec- 

 trophotometrically that 1 mol ADH binds 2 mols of DPNH. 

 For the sake of convenience we shall denote V2 ADH as ADH'. 



Furthermore, by using rapid spectrophotometry Theorell 

 and Chance (24) could determine the velocity of the band shift. 



J/o 320 330 J-^O 350 



Fig. 5. Millimolar extinction of free and ADH-bound 

 DPNH. Ordinate: e cm.-i X mM-i. (From ref. 23.) 



and thus obtained the value 4 X 10^ M~^ sec. "^ for the velocity 

 constant (ki) of the combination between DPNH and ADH. 

 In the equilibrium 



[DPNH][CH3CHO][H+] 

 [DPNJlCaHeOH] 



Racker (15), using small amounts of yeast ADH, had found K to 

 be constant and independent of pH, as required by the formula. 

 The value for K was close to 10~". 



Since iT is a function of purely thermodynamic parameters, 

 it was no surprise that we found the same value with low concen- 

 trations of liver ADH. But in the liver the concentration of the 

 enzyme (^ 1 g. per kilogram wet weight) is not at all low enough 

 to be negligible. 



294 



