Interactions between carbonic anhydrase and some decarboxylating enzymes as studied by a new bioelectrochemical approach

This work presents the results of a study, carried out by recently develope d amperometric bioelectrodes, on the interactions between carbonic anhydras e (CA) and the decarboxylating enzymes arginine decarboxylase (ADC), L-lysi ne decarboxylase (LDC), and L-ornithine decarboxylase (ODC). These are all pyridoxal-phosphate dependent enzymes and catalyze the decarboxylation reac tion of the respective amino acids, to give carbon dioxide and the correspo nding diamine (agmatine, cadaverine, and putrescine, respectively). The rat e of each decarboxylase catalyzed reaction was measured by monitoring the p roduction of the respective diamine by a plant tissue diamino oxidase (DAO) based bioelectrode. DAO is the enzyme which catalyzes the oxidation of agm atine, cadaverine, and putrescine with the production of NH3 and H2O2. DAO- based bioelectrodes consist of an amperometric H2O2 electrode, coupled to t he biocatalytic membrane formed by a whole plant tissue (lentil cotyledon) containing the enzyme DAO, immobilized on a dialysis membrane by polyazetid ine prepolymer (PAP). The bioelectrodes were calibrated and characterized i n standard solutions of agmatine, cadaverine, and putrescine. Kinetic studi es to measure decarboxylase activity were performed in the presence of diff erent concentrations of ADC, LDC, and ODC, resulting in a lowest detection limit of 10, 25, and 10 U l(-1) respectively. The effect of bovine CA II (b CAII) was evaluated in the presence of 500 U l(-1) of each decarboxylase, s howing a marked increase of the rate of the decarboxylation reaction. These results suggest that (i) CA can be used to enhance the performance of deca rboxylase-based biosensors, and (ii) it possibly plays further physiologica l roles, acting synergistically, at specific cellular and subcellular sites , with low-activity decarboxylating enzymes. (C) 1999 Elsevier Science S.A. All rights reserved.