78 3. KINETICS OF ENZYME INHIBITION 



moles, and it is likely that the system will be in zone B or C depending on 

 the value of K,. Prediction of competitive inhibition kinetics under such 

 circumstances is extremely difficult since we never accurately know the 

 localized substrate concentration; it is likely that competitive inhibition as 

 it is usually formulated is uncommon in these cases. 



Role of Extraneous Substances in Mutual Depletion Systems 



In the systems so far discussed it has been assumed that one is dealing 

 with a pure enzyme or, if not, that other material does not bind the inhibi- 

 tor. In many enzyme preparations there are substances that can bind cer- 

 tain inhibitors as well, or even better, than the enzyme; protein impurities 

 can usually bind heavy metals, arsenicals, and protein reactants extensively. 

 Now, binding of inhibitors by such extra-enzyme material will cause the 

 system to approach or enter zones B or C, because it makes no difference 

 what binds the inhibitor and reduces its free concentration. Actually in 

 systems containing binding components other than the enzyme, it is not the 

 value of (E/) that determines the zone but a sum of such terms including 

 each binding substance, or approximately considered the total concentra- 

 tion of binding sites divided by an average dissociation constant. This would 

 apply particularly to impure enzyme preparations, mitochondria, homogen- 

 ates, and cell suspensions. If we designate the specific concentration of all 

 binding sites by (X/) it is to this that the boundary conditions really apply 

 and (X/) may well be 100-1000 times larger than (E/). Conversely, in such 

 situations, an estimation of enzyme concentration, as discussed above, may 

 be erroneous. This will be taken up in more detail in Chapter 15. 



PSEUDO-IRREVERSIBLE INHIBITION 



Ackermann and Potter (1949) defined pseudo-irreversible inhibitors as 

 those that have such a high affinity for the enzyme that they give the 

 appearance of irreversibility. The inhibition may be competitive or non- 

 competitive, but if it is truly the former, it may not be obvious since little 

 change in enzyme activity will be observed upon adding the substrate to 

 the inhibited enzyme, due to the slow dissociation of inhibitor, or the in- 

 bitor is bound so much more tightly to the enzyme than is the substrate 

 that com])etition at equilibrium cannot be demonstrated. This is really 

 another approach to systems in zones B or C and the equations given by 

 Ackermann and Potter are identical to those of Straus and Goldstein for 

 comparable situations; thus the expression for the inhibited rate given by 

 Ackermann and Potter, i\ = A;(E^) — ^'(I,), is equivalent to an inhibition 

 of i = (I^)/(E^), since v = ^(E^). However, a short exposition of this ap- 

 proach will serve to introduce some further practical applications for such 

 inhibitors. 



