The pH-dependence of enzymic ester hydrolysis.

The esterases of animal tissues have generally been characterized by their substrate specificity and by their behaviour towards various inhibitors. The irreversible inhibition by organic phosphates, which is due to phosphorylation of the active group (Burgen, 1949; Wilson & Bergmann, 1950a; Aldridge, 1953d), is a common property of most esterases, an interesting exception being the Aesterase of Aldridge (1953a, b). This general behaviour suggests a similar structure of most of the esteratic groups responsible for the hydrolytic reaction. True cholinesterase has been characterized, in addition to the properties mentioned, by the dissociation constants of its esteratic site (Wilson & Bergmann, 1950 b). However, the negative charges present in the active centre of this enzyme are also pH-dependent and thus interpretation of the experimental data obtained with acetylcholine (ACh) as substrate is difficult (Bergmann & Shiimoni, 1952 a). This difficulty can be met by studying the influence of pH changes on the enzymic hydrolysis of uncharged esters, since in such a system all variations of activity can be ascribed solely to structural changes of the esteratic site. In the present investigation, such procedures have been applied to true and pseudocholinesterase and to an unspecific liver esterase. From these experiments certain conclusions about the chemical structure of the esteratic sites can be drawn, and a rational interpretation can be derived for the bellshaped curves relating hydrolytic rates to substrate concentration which have been found not only in the system true cholinesterase-acetylcholine, but with other esterases and a variety of uncharged substrates (Bergmann & Shimoni, 1953). In addition, the dissociation constants of the anionic sites in both types of cholinesterases have been determined and a tentative explanation of the curious values obtained has been advanced. Part of the results described in this paper have been communicated at the Meeting of the Faraday Society held at Oxford in August 1955 (Bergmann, 1955).