198 6. INTERACTIONS OF INHIBITORS WITH ENZYMES 



occurring during the reaction (Laidler, 1955 a). Thus enzymic hydrolysis 

 can occur following the multipoint binding of the substrate by a " push- 

 pull " acid-base catalysis exerted by two groups, one acidic and one basic. 

 That there are binding groups determining the orientation and specificity 

 and also groups activating a labile bond has been postulated for cliymo- 

 trypsin on the basis of the entropy differences observed between optical 

 isomers of the substrates (Vaslow and Doherty, 1953). This concept is 

 inherent in the mechanisms ascribed to several enzymes, such as succinic 

 dehydrogenase, fumarase, acetylcholinesterase, carboxypeptidase, proli- 

 dase, and others. In more complex reactions, two or more of such sites 

 may cooperate, especially in transfer processes; such is evident in many 

 oxidation-reduction reactions but may also occur in transfer associated 

 with phosphatases and other hydrolytic enzymes, the acceptor being either 

 water for hydrolysis or some other molecule for transfer, either acceptor 

 being bound to a specific site adjacent to the donor site (Morton, 1955). 

 A metal ion, complexed to the protein, may serve as a site for either binding 

 or reaction or both but full catalytic effectiveness is only achieved in con- 

 junction with other sites. The transphosphorylation ATP + P^^ -> ADP 

 _l_ pp32 ig catalyzed by various metal ions (Mn"^"^, Ca"^"^, Cd"^"*") in the ab- 

 sence of any enzyme, but the concentration of substrates must be about 

 fifty-fold greater than for the enzymic reaction, indicating that the addi- 

 tional groups on the enzyme increase the efficiency of the reaction by orien- 

 tation in the proper position (Lowenstein, 1958). Spatial coordination 

 of groups within active sites, and of sites within an active center, is of- 

 ten necessary for the catalysis; as a consequence, disturbances in this struc- 

 tural pattern can inhibit such enzymes. 



Despite the fact that most enzymes are globular or ellipsoidal, the sur- 

 face should not be considered as even approximately smooth but, in the 

 most interesting regions, perhaps more like a holly bush. At least the various 

 protrusions, indentations, and crevices that undoubtedly occur are occa- 

 sionally important in the configuration of the active site according to sev- 

 eral recent speculations on the ES complex as in Figs. 6-1 and 6-2. The 

 advantage to the " action in a hole " concept is that more groups can be 

 brought to bear on the reactants and attack can be made from opposite 

 sides. It must be admitted that there is essentially no direct evidence for 

 such sites but in peroxidase and catalase the presence of the heme in a 

 protein crevice is made likely by inhibition and ionization studies (George 

 and Lyster, 1957). Such crevices may exist between helices. An active site 

 may be situated in a helix but it is more likely that it comprises portions 

 of the polypeptide chain in more extended configurations, or combinations 

 of the two structures; in any event it is evident that tertiary protein struc- 

 ture is of significance in enzymes and that alterations of this structure 

 even at some distance from the active site, can modify the catalysis. 



