Reverse Ordered Sequential Mechanism for Lactoperoxidase with Inhibition by Hydrogen Peroxide.

Kellye Cupp-Sutton,Michael T Ashby

doi:10.3390/antiox10111646

Abstract

Lactoperoxidase (LPO, FeIII in its resting state in the absence of substrates)—an enzyme secreted from human mammary, salivary, and other mucosal glands—catalyzes the oxidation of thiocyanate (SCN−) by hydrogen peroxide (H2O2) to produce hypothiocyanite (OSCN−), which functions as an antimicrobial agent. The accepted catalytic mechanism, called the halogen cycle, comprises a two-electron oxidation of LPO by H2O2 to produce oxoiron(IV) radicals, followed by O-atom transfer to SCN−. However, the mechanism does not explain biphasic kinetics and inhibition by H2O2 at low concentration of reducing substrate, conditions that may be biologically relevant. We propose an ordered sequential mechanism in which the order of substrate binding is reversed, first SCN− and then H2O2. The sequence of substrate binding that is described by the halogen cycle mechanism is actually inhibitory.

Highlights

Mechanism for Lactoperoxidase withSecretory proteins play essential roles in human host defense
We have identified the LPO-H2O2 bound species as Comof 14 pound I* (Cpd I*-[FeIV-OH, aa+.], an oxoiron(IV) radical like Cpd I, albeit protonated2[6,7,8], but with the organic radical relocated from the porphyrin to an amino acid)
After a model is presented that describes the kinetics, we discuss the possible structures of species in the context of extra-kinetic data and the literature

Summary

Introduction

Mechanism for Lactoperoxidase withSecretory proteins play essential roles in human host defense. Perhaps the most widely studied secretory protein is lactoperoxidase (LPO, EC 1.11.1.7) [1]. The accepted catalytic mechanism, a two-electron oxidation of native LPO-[FeIII ] (Figure 1A, Equation (1)) followed by O-atom transfer from the enzyme to the substrate (Figure 1A, Equation (2)), is often referred to as the “halogen cycle”. According to this widely accepted mechanism, substrate selectivity is determined by the rate of the reaction of Cpd I with potential substrates [3], which in turn is a function of the rate constants for the second step of the halogen cycle mechanism and the concentrations of the substrates in various physiologic fluids [4]. The antimicrobial activity of the LPO system is attributed to hypothiocyanite (OSCN− ) [5]

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Conclusion