Complexes between m-chloroperoxybenzoic acid and horseradish peroxidase compounds I and II: Implications for the kinetics of enzyme inactivation

Abstract

In summary, the complexes between HRP-C compounds I or II, and mCPBA have been identified as being functionally equivalent to enzyme that has been inactivated due to the activity of this substrate as a mechanism-based inactivating agent, in as much that the affinity of mCPBA for HRP-C is high enough that the H2O2 used in assays with the donor substrate ABTS cannot displace mCPBA unless it is present in very large excess. These observations provide an explanation of the discrepancy between the time dependent falls in activity with ABTS and simulations of the reaction mechanism (Figure 1) performed using accurately determined rate constants from single turnover stopped-flow experiments. The data provide a useful addition to our knowledge of the behaviour of this widely studied enzyme that nevertheless continues to offer further opportunities for research. They also indicate that whilst steady state and transient state kinetic data may produce apparently consistent results it is important to bear in mind other circumstances such as the assay conditions being used.