Eukaryotic extracellular catalase-peroxidase from Magnaporthe grisea - Biophysical/chemical characterization of the first representative from a novel phytopathogenic KatG group |
| |
Authors: | Zámocký Marcel Droghetti Enrica Bellei Marzia Gasselhuber Bernhard Pabst Martin Furtmüller Paul G Battistuzzi Gianantonio Smulevich Giulietta Obinger Christian |
| |
Institution: | Division of Biochemistry, Department of Chemistry, Vienna Institute of Biotechnology at BOKU - University of Natural Resources and Life Sciences, Muthgasse 18, A-1190 Vienna, Austria. marcel.zamocky@boku.ac.at |
| |
Abstract: | All phytopathogenic fungi have two catalase–peroxidase paralogues located either intracellularly (KatG1) or extracellularly (KatG2). Here, for the first time a secreted bifunctional, homodimeric catalase–peroxidase (KatG2 from the rice blast fungus Magnaporthe grisea) has been produced heterologously with almost 100% heme occupancy and comprehensively investigated by using a broad set of methods including UV–Vis, ECD and resonance Raman spectroscopy (RR), thin-layer spectroelectrochemistry, mass spectrometry, steady-state &; presteady-state spectroscopy. RR spectroscopy reveals that MagKatG2 shows a unique mixed-spin state, non-planar heme b, and a proximal histidine with pronounced imidazolate character. At pH 7.0 and 25 °C, the standard reduction potential E°′ of the Fe(III)/Fe(II) couple for the high-spin native protein was found to fall in the range typical for the KatG family. Binding of cyanide was relatively slow at pH 7.0 and 25 °C and with a Kd value significantly higher than for the intracellular counterpart. Demonstrated by mass spectrometry MagKatG2 has the typical Trp118-Tyr251-Met277 adduct that is essential for its predominantly catalase activity at the unique acidic pH optimum. In addition, MagKatG2 acts as a versatile peroxidase using both one- and two-electron donors. Based on these data, structure–function relationships of extracellular eukaryotic KatGs are discussed with respect to intracellular KatGs and possible role(s) in host–pathogen interaction. |
| |
Keywords: | Extracellular catalase–peroxidase Peroxidases–catalase superfamily Phytopathogen Oxidative stress Resonance Raman spectroscopy Reduction potential |
本文献已被 ScienceDirect PubMed 等数据库收录! |
|