EFFECTS OF THE DIELECTRIC CONSTANT 809 



enzyme inhibition unless there is good evidence that one type of intermo- 

 lecular force is preponderant. 



Lowering the dielectric constant should increase the rates of reaction 

 between oppositely charged ions or between any molecules where electro- 

 static attraction of any type is important in the formation of the activated 

 complex. This has been tested on many nonenzymic systems, especially 

 the displacement reaction between thiosulfate and bromoacetate (where 

 the rate constant decreases with decreasing dielectric constant since these 

 are both anions) and the reactions of various substances with hydrogen or 

 hydroxyl ions, such as the proton-catalyzed splitting of the azodicarbonate 

 ion (where a marked rise in the rate constant with decrease in dielectric 

 constant is observed), or the reaction of bromophenol blue with hydroxyl 

 ions. Qualitative correspondence to the theoretical expectations is always 

 found but quantitative differences occasionally occur and these may be 

 due to hydration changes in solvent mixtures and to the use of the bulk 

 dielectric constants. 



Experimental Studies of Enzyme Reactions 



The effect of changes in the dielectric constant on the rates of enzyme 

 reactions can be attributed to alterations in the affinity of the enzyme for 

 the substrate, or to alterations in the rate of breakdown of the ES complex, 

 or to both. In other words, there may be changes in A',, ii ,, or K„j. The over- 

 all effect on the rate may be complex. For example, in the reaction of a 

 negatively charged substrate with a positively charged active site: 



S- -f- E+ — ES ^ E+ + P- 



in which the product is also negatively charged, a lowering of the dielectric 

 constant will decrease both K, and A'a, the former tending to increase and 

 the latter tending to decrease the rate. The magnitude of the effect will, 

 of course, depend also on the substrate concentration relative to K, or 

 K,,,, since only changes in A-., will be important when the enzyme is substrate- 

 saturated but changes in K, or /iT,,, will be increasingly significant as the 

 substrate concentration is reduced. 



(a) a-Chymotrypsin. The addition of methanol up to concentrations 

 around 30% (D = 64.3) progressively slows the rates of hydrolysis of a 

 number of synthetic substrates. The logarithm of the proteolytic coefficient 

 for the hydrolysis of glycyl-L-tyrosinamide and benzoyl-L-tyrosine ethyl 

 ester decreases linearly with the methanol concentration (Kaufman et al., 

 1949). With acetyl-L-tyrosinamide as the substrate, it was subsequently 

 shown that the rate constant k^ is unaltered while K,„ increases from 0.0326 

 to 0.0806, when 20.8% methanol is present (Kaufman and Neurath, 1949). 



