Proton Equilibria in the Binding of Zn²⁺ and of Methylmercuric Iodide to Papain

Abstract

The proton liberation on the binding of zinc chloride and methylmercuric iodide to the (essential) thiol group of papain has been examined as a function of pH. This was carried out by (a) direct titration of the protons on the addition of the metal compound to active papain and (b) measurement of the extent of inhibition of enzyme activity by the metal compound as a function of pH.It was found that in the neutral pH range the thiol group or the neighbouring imidazole group in the free enzyme carries one proton, at low pH both groups do so, whereas at high pH neither group carries a proton. The pK values of the free enzyme that govern the proton release, 4.2 and 8.5, correspond to those that govern overall activity. Both from the experiments with methylmercuric iodide and from fluorescence measurements of methylmercuric papain, it was established that the imidazole group in the latter compound exhibits a pK of 5.4. Taking recent data into account, it was considered that the ion pair of thiolate anion and imidazolium cation, proposed by Polgar, is the best approximation to describe the charge distribution in the active centre and to explain the reaction mechanism.In the trough of the active site of papain the essential thiol group of cysteine-25 and the imidazole group of histidine-159 are located within 1 nm of each other [l].

Removal of mercury from its aqueous solution using charcoal-immobilized papain (CIP)

“…In the present article to assess the efficiency of the process, the desorption of mercury from CIP-mercury complex has also been investigated at different pH values. It has been seen that low pH facilitates the desorption process which is in agreement with the observation made by Sluyterman and Wijdenes [12]. Therefore, it can be stated that papain, having the characteristics of metal binding due to presence of four sulfhydryl groups in its active sites, can be used to modify the free matter space of activated charcoal, a micro-porous solid matrix, and thereby increasing the number of active sites of charcoal for the removal of mercury from its aqueous solution.…”

“…The inhibitory effect of mercury ion to enzymatic action of papain proves the binding capability of mercury ion with papain. Moreover, with a high affinity towards organic compounds mercury has the possibility to bind with other amino acids [11][12][13]. Hence, papain, immobilized on suitable matrix, can be used in polishing treatment of industrial wastes for removal of mercury.…”

Identification of the active site cysteine and of the disulfide bonds in the N‐terminal part of the molecule of bovine spleen cathepsin B

“…Papain behaves similarly [24] and it has been postulated that the Cys-22-Cys-63 bridge is reduced. Therefore, it is possible that it is the interchain S-Sbridge of CB which is preferentially reduced.…”

Removal of mercury (II) from aqueous solution using papain immobilized on alginate bead: Optimization of immobilization condition and modeling of removal study

“…Binding of metal ion with the dissociated state E À is ionic in nature and therefore expected to be fast. According to Sluyterman and Wijdenes (1976), the reaction is reversible. These are the considerations for assuming the equilibrium reaction (2), which represents binding of metal ion to E À and their dissociation, as part of the proposed mechanism.…”