Characterization of a catalase-peroxidase from the hyperthermophilic archaeon Archaeoglobus fulgidus

A putative perA gene from Archaeoglobus fulgidus was cloned and expressed in Escherichia coli BL21(DE3), and the recombinant catalase-peroxidase was purified to homogeneity. The enzyme is a homodimer with a subunit molecular mass of 85 kDa. UV-visible spectroscopic analysis indicated the presence of...

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Veröffentlicht in:Extremophiles : life under extreme conditions 2001-10, Vol.5 (5), p.323-332
Hauptverfasser: Kengen, S W, Bikker, F J, Hagen, W R, de Vos, W M, van der Oost, J
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container_title Extremophiles : life under extreme conditions
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creator Kengen, S W
Bikker, F J
Hagen, W R
de Vos, W M
van der Oost, J
description A putative perA gene from Archaeoglobus fulgidus was cloned and expressed in Escherichia coli BL21(DE3), and the recombinant catalase-peroxidase was purified to homogeneity. The enzyme is a homodimer with a subunit molecular mass of 85 kDa. UV-visible spectroscopic analysis indicated the presence of protoheme IX as a prosthetic group (ferric heme), in a stoichiometry of 0.25 heme per subunit. Electron paramagnetic resonance analysis confirmed the presence of ferric heme and identified the proximal axial ligand as a histidine. The enzyme showed both catalase and peroxidase activity with pH optima of 6.0 and 4.5, respectively. Optimal temperatures of 70 degrees C and 80 degrees C were found for the catalase and peroxidase activity, respectively. The catalase activity strongly exceeded the peroxidase activity, with Vmax values of 9600 and 36 U mg(-1), respectively. Km values for H2O2 of 8.6 and 0.85 mM were found for catalase and peroxidase, respectively. Common heme inhibitors such as cyanide, azide, and hydroxylamine inhibited peroxidase activity. However, unlike all other catalase-peroxidases, the enzyme was also inhibited by 3-amino-1,2,4-triazole. Although the enzyme exhibited a high thermostability, rapid inactivation occurred in the presence of H2O2, with half-life values of less than 1 min. This is the first catalase-peroxidase characterized from a hyperthermophilic microorganism.
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subjects Archaeoglobus fulgidus - enzymology
Archaeoglobus fulgidus - genetics
Catalase - antagonists & inhibitors
Catalase - chemistry
Catalase - genetics
Catalase - metabolism
Cloning, Molecular
Electron Spin Resonance Spectroscopy
Enzyme Inhibitors - pharmacology
Genes, Archaeal
Hot Temperature
Kinetics
Peroxidases - antagonists & inhibitors
Peroxidases - chemistry
Peroxidases - genetics
Peroxidases - metabolism
Phylogeny
Recombinant Proteins - chemistry
Recombinant Proteins - genetics
Recombinant Proteins - metabolism
Sequence Analysis, Protein
Space life sciences
Spectrophotometry
title Characterization of a catalase-peroxidase from the hyperthermophilic archaeon Archaeoglobus fulgidus
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