How Does Replacement of the Axial Histidine Ligand in Cytochrome c Peroxidase by Nδ-Methyl Histidine Affect Its Properties and Functions? A Computational Study.

Calvin W Z Lee,Sam P De Visser,M Qadri E Mubarak,Anthony P Green

doi:10.3390/ijms21197133

Calvin W Z Lee, Sam P De Visser + Show 2 more

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https://doi.org/10.3390/ijms21197133

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Abstract

Heme peroxidases have important functions in nature related to the detoxification of H2O2. They generally undergo a catalytic cycle where, in the first stage, the iron(III)–heme–H2O2 complex is converted into an iron(IV)–oxo–heme cation radical species called Compound I. Cytochrome c peroxidase Compound I has a unique electronic configuration among heme enzymes where a metal-based biradical is coupled to a protein radical on a nearby Trp residue. Recent work using the engineered Nδ-methyl histidine-ligated cytochrome c peroxidase highlighted changes in spectroscopic and catalytic properties upon axial ligand substitution. To understand the axial ligand effect on structure and reactivity of peroxidases and their axially Nδ-methyl histidine engineered forms, we did a computational study. We created active site cluster models of various sizes as mimics of horseradish peroxidase and cytochrome c peroxidase Compound I. Subsequently, we performed density functional theory studies on the structure and reactivity of these complexes with a model substrate (styrene). Thus, the work shows that the Nδ-methyl histidine group has little effect on the electronic configuration and structure of Compound I and little changes in bond lengths and the same orbital occupation is obtained. However, the Nδ-methyl histidine modification impacts electron transfer processes due to a change in the reduction potential and thereby influences reactivity patterns for oxygen atom transfer. As such, the substitution of the axial histidine by Nδ-methyl histidine in peroxidases slows down oxygen atom transfer to substrates and makes Compound I a weaker oxidant. These studies are in line with experimental work on Nδ-methyl histidine-ligated cytochrome c peroxidases and highlight how the hydrogen bonding network in the second coordination sphere has a major impact on the function and properties of the enzyme.

Highlights

Heme peroxidases perform a wide range of functions in biology, ranging from defense mechanisms against infection to hormone biosynthesis in the human body [1,2,3,4,5,6,7,8,9]
horseradish peroxidase (HRP) Compound I (Cpd I) was modelled as a minimal model with the heme group approximated as protoporphyrin IX, with all side chains replaced by hydrogen atoms, an oxo in the distal position and with imidazole as an axial ligand bound to iron
Model B included a nearby K+ ion to obtain an electronic configuration for Cpd I that reflects the experimental electron paramagnetic resonance (EPR) data for cytochrome c peroxidase (CcP)

Summary

Introduction

Heme peroxidases perform a wide range of functions in biology, ranging from defense mechanisms against infection to hormone biosynthesis in the human body [1,2,3,4,5,6,7,8,9]. The peroxidases undergo a catalytic cycle starting from a resting state, i.e., a water-bound heme group, where the water ligand is substituted by H2O2. This H2O2 binding triggers a proton relay that converts the H2O2-bound heme into water and an iron(IV)–oxo heme cation radical intermediate, called Compound I (Cpd I). The difference in the protein binding pattern between the peroxidases and mono-oxygenases generally has an effect on the properties of the iron/heme co-factor as the more electron donating cysteinate ligand induces a stronger “push” effect compared with the neutral histidine-ligated system [25,26,27]. Mutations of the axial ligand or its direct environment affect the structure and catalytic ability of the protein [28,29,30,31]

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