76 3. KINETICS OF ENZYME INHIBITION 



as i = (I^)/(E^). As long as the system is in zone C, most of the inhibitor 

 is bound to the enzyme and the fraction of inactive enzyme is independent 

 of the potency of the inhibitor. It is obvious that no constants may be cal- 

 culated from zone C kinetics. 



Dilution of Enzyme-Inhibitor Systems 



The characteristic changes in inhibition brought about by dilution were 

 used by Straus and Goldstein as criteria for zone behavior. Two types of 

 dilution must be distinguished: {a) only the enzyme is diluted and the inhi- 

 bitor is added at the same concentration and {b) the enzyme-inhibitor mix- 

 ture is diluted. Enzyme dilution alone has no effect on the inhibition in zone 

 A and increases inhibition in zones B and C, or may translate the system from 

 zone C into B or from zone B into A; this is evident from the dependence of 

 zone behavior on (E/). In the second situation where both enzyme and in- 

 hibitor are diluted (the " dilution effect " of Straus and Goldstein), equally 

 simple relations are observed for noncompetitive inhibition. In zone A 

 dilution leads to less inhibition simply because the inhibitor concentration 

 becomes smaller and the enzyme concentration is of no importance; in zone 

 C both (I/) and (E/) change equally and there is no effect on inhibition. In 

 zone B the situation is somewhat more complex because both (I/) and (E/) 

 are involved in determining the inhibition. Dilution will produce a decrease 

 in inhibition, due to reduction of (I/), but the decrease will be less than in 

 zone A because the reduction of (E/) will tend to increase the inhibition 

 somewhat; thus in zone B the change of inhibition on dilution depends on 

 the change in (E/) whereas in zone A it does not. Dilution of competitively 

 inhibited systems also involves decrease in (S/). In zone A^A^ there will 

 be no change in inhibition if the assumptions leading to Eq. 3-50 are valid, 

 since both (I/) and (S/) change equally; however, from the more general 

 Eq. 3-47 it is clear that inhibition will increase. In zones A,B, and A^Cj 

 there will also be no change in inhibition on dilution if the simplifications 

 giving Eqs. 3-52 and 3-54 are valid, but if the more general Eq. 3-48 must 

 be used, there is a dilution effect which also depends on (E/) in zone A,B,. 

 In case the system is in zones B, or C^ the dilution effect is complex and 

 depends generally on all four specific concentrations, (S/), (I/), (E,'), and 

 (E/), as is evident from Eq. 3-45. The complex equations for the magnitude 

 of the dilution effect in noncompetitive systems are given by Straus and 

 Goldstein and a graphical presentation for competitive systems by Gold- 

 stein, together with the effects of dilution on cholinesterase inhibition. 



Noncellular preparations usually represent enzymes diluted from the 

 concentration existing in the cell; this is dilution of the first type, i.e., of 

 enzyme only. Sometimes a tissue is exposed to an inhibitor and the inhi- 

 bited enzyme is subsequently extracted for the purpose of determining the 

 extent of inhibition occurring in the tissue; this is dilution of the second type. 



