Synthesis and kinetic evaluation of a trifunctional enzyme mimic with a dimanganese active centre

Abstract

Glutathione peroxidase (GPX), superoxide dismutase (SOD) and catalase (CAT) play crucial roles in the metabolism and homeostasis of reactive oxygen species (ROS) in living organisms. From examination of the steady state and pre-steady state kinetic behavior of natural GPX it was found that, in contrast to accepted theories, the affinity of the enzyme for H2O2 rather than reduced glutathione (GSH) most significantly affects its kinetic behavior. Consequently, an enzyme mimic was produced with a similar affinity for the substrate H2O2. A salicylaldehyde Schiff base containing a dimanganese centre was selected as a precursor, because it has high H2O2-binding affinity for such a relatively small molecule and similar catalytic activity to that of SOD and CAT. Selenium was also incorporated into the catalytic center to provide activity similar to that of GPX, and thus trifunctional enzymatic activity. The KmH2O2 of the mimic (7.32×10-2mM) was found quite close to that of natural enzyme (1.0×10-2mM), indicating that the affinity of the mimic to H2O2 was successfully increased to approach natural GPX. The steady state kinetic performance of the enzyme mimic showed that the ratio between kcat/KmGSH and kcat/ KmH2O2 was quite similar to that of native GPX, indicating that the Mn(III)2(L-Se-SO3Na) had the same selectivity for both substrates GSH and H2O2 as native GPX, which put it among the best existing GPX mimics. Moreover, the new mimic was confirmed to strongly inhibit lipid peroxidation and mitochondrial swelling, probably due to the synergism between the three antioxidant enzymatic activities.