The paradoxical influence of the pKa on the reactivity of thiols and its biological relevance

Abstract

Background. Not much is known about the factors that influence thiol reactivity. In this paper we will investigate the paradoxical influence of the acid dissociation constant (pKa). The pKa influences the deprotonation of a compound. This deprotonation gives thiols a negative charge and therefore increases reactivity. Compounds with a low pKa have a higher percentage of deprotonation, this will lead to a higher reactivity. On the other side, thiols with a higher pKa have a higher electron density, and are therefore also more reactive. We want to recognize this paradox and investigate what the exact contribution of these two paradoxical mechanisms is with respect to the reactivity of thiols. Furthermore we will discuss the biological importance of this study with respect to the anti-oxidant system and drug conjugation. Methods. The reactivity of the thiols was determined by calculating the rate constant of these compounds. This was done for different thiols in reaction with 1-chloro-2,4- dinitrobenzene. To do this we created a pseudo first order reaction by adding an excess of CDNB. The thiols we used in this reaction were glutathione, 5-thio-2-nitrobenzoic acid, 2-mercaptoethanol and 2-mercaptobenzothiazole. The reaction rates were measured, using a spectrophotometer and eventually the reaction constant was calculated and transformed into the reactivity of the thiolate ion. Results. The reaction constants from the different thiols are dependent on the fraction thiolate ion. This was converted to the reactivity of the thiolate ion. The reactivity of thiolate ion decreases as pKa value increases. Conclusion. There is indeed a paradoxical influence of the pKa value on the reactivity of thiols. Increases in pKa lead to decreases in reactivity of the thiolate ion but to increases in the fraction of the thiolate ion. Eventually there was no mechanism that had the biggest contribution to the reactivity for all conditions. Apparently this is dependent on the pH. We advice drug researchers to keep this paradox in mind in order to estimate the reactivity of thiols.