800 15. EFFECTS OF VARIOUS FACTORS ON INHIBITION 



enzyme interactions and the nature of the forces between inhibitors and 

 active sites. 



Water has a high dielectric constant and this reduces appreciably the 

 interactions between ions, ions and dipoles, and dipoles. If the dielectric 

 constant is lowered, one would expect that these electrostatic interactions 

 would become greater. Not only would two oppositely charged ions associate 

 more readily but repulsion between similarly charged ions would increase. 

 Electrokinetic or dispersion forces would not be significantly altered. Thus 

 the formation of most enzyme-inhibitor complexes would occur at a faster 

 rate upon reduction of the dielectric constant and the binding energy might 

 be increased. The variation of the affinity of the enzyme for the inhibitor, 

 or of the inhibitor constant K^, with the dielectric constant may provide 

 some information on the nature of the interaction. 



Methods for Reduction of the Dieletric Constant 



The dielectric constant of aqueous media may be decreased by the addi- 

 tion of various organic liquids that are miscible with water. The most com- 

 monly used substances for this purpose are dioxane, methanol, ethanol, 

 w-propanol, isopropanol, methyl ethyl ketone, acetone, acetonitrile, dimeth- 

 ylacetamide, ^e/^butanol, ethylene glycol, and glycerol. Although die- 

 lectric constants from 2 to 78 may be obtained by such mixtures, one is 

 limited in enzyme work to solutions containing not over 50% of the organic 

 solvent and thus the dielectric constant can usually not be reduced below 

 35. Tables giving the dielectric constants of mixtures of water with various 

 solvents are available (Harned and Owen, 1958, p. 161). Plots for three 

 solvents are shown in Fig. 15-15. Over the range used in enzyme study, 

 the relationship between dielectric constant and solvent concentration 

 is linear. One of the most useful organic solvents is dioxane; it is com- 

 pletely miscible with water, it is nonreactive and electrically neutral, 

 and its low dielectric constant allows rates to be studied over a relatively 

 wide range. It must be remembered that the dielectric constant varies with 

 the temperature. Most of the values given for dielectric mixtures in the 

 chemical literature are for 25" and if the enzyme reaction is run at more 

 physiological temperatures the appropriate correction should be made. 

 Over a range of dioxane concentration from 10% to 70%, the temperature 

 correction may be obtained from the approximate relationship: 



— = _ 0.0023 (15-57) 



Thus in 30% dioxane, Z)(25o) = 51.9 and D(37o) = 48.6. 



