Abstract
Reconstructing the phylogenetic relationships of the main evolutionary lines of the mammalian peroxidases lactoperoxidase and myeloperoxidase revealed the presence of novel bacterial heme peroxidase subfamilies. Here, for the first time, an ancestral bacterial heme peroxidase is shown to possess a very high bromide oxidation activity (besides conventional peroxidase activity). The recombinant protein allowed monitoring of the autocatalytic peroxide-driven formation of covalent heme to protein bonds. Thereby, the high spin ferric rhombic heme spectrum became similar to lactoperoxidase, the standard reduction potential of the Fe(III)/Fe(II) couple shifted to more positive values (-145 ± 10 mV at pH 7), and the conformational and thermal stability of the protein increased significantly. We discuss structure-function relationships of this new peroxidase in relation to its mammalian counterparts and ask for its putative physiological role.
Highlights
First analysis was made of bacterial ancestor of peroxidases from mammalian innate immune system
Reconstructing the phylogenetic relationships of the main evolutionary lines of the mammalian peroxidases lactoperoxidase and myeloperoxidase revealed the presence of novel bacterial heme peroxidase subfamilies
Thereby, the high spin ferric rhombic heme spectrum became similar to lactoperoxidase, the standard reduction potential of the Fe(III)/Fe(II) couple shifted to more positive values (؊145 ؎ 10 mV at pH 7), and the conformational and thermal stability of the protein increased significantly
Summary
First analysis was made of bacterial ancestor of peroxidases from mammalian innate immune system. PCC 8106 as there seems to be a direct lineage from this bacterial oxidoreductase to the mammalian peroxidases (Fig. 1) This first investigated member of the peroxidockerin clade could be produced in Escherichia coli with almost 100% heme occupancy. The recombinant enzyme for the first time allowed monitoring of the autocatalytic formation of covalent heme to protein bonds by a broad set of techniques, including UV-visible, ECD, and EPR spectroscopy, mass spectrometry, spectroelectrochemistry, and differential scanning calorimetry These findings are discussed with respect to published structure-function relationships of LPO, EPO, and MPO, the putative physiological role of these metalloenzymes in bacteria, and their potential application in biotechnology
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