Crystal Structure of Dihydro-Heme d1 Dehydrogenase NirN from Pseudomonas aeruginosa Reveals Amino Acid Residues Essential for Catalysis |
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| Affiliation: | 1. Department of Structure and Function of Proteins, Helmholtz Centre for Infection Research, Inhoffenstrasse 7, 38124 Braunschweig, Germany;2. Institute of Biochemistry, Leipzig University,Brüderstrasse 34, 04103 Leipzig, Germany;3. Department of Environmental Microbiology, Helmholtz Centre for Environmental Research, Permoserstrasse 15, 04318 Leipzig, Germany;4. Institute of Pharmaceutical Sciences, University of Freiburg, Stefan-Meier-Str.19, 79104 Freiburg, Germany;5. Institute for Biochemistry, Biotechnology and Bioinformatics, Technische Universität Braunschweig, 38106 Braunschweig, Germany;1. Institute for Quantum Life Science, National Institutes for Quantum and Radiological Science and Technology, 2-4 Shirakata, Tokai, Ibaraki 319-1106, Japan;2. Nagoya University Synchrotron Radiation Research Center, Furo-cho, Chikusa-ku, Nagoya, Aichi 464-0965, Japan;3. Structural Biology Research Center, Garduate School of Science, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Aichi 464-8601, Japan;4. Neutron Science Section, Materials and Life Science Division, J-PARC Center, Japan Atomic Energy Agency, 2-4 Shirakata, Tokai, Ibaraki 319-1195, Japan;1. Center for Structural Systems Biology, Helmholtz Centre for Infection Research, Department of Structural Infection Biology, Notkestraße 85, 22607 Hamburg, Germany;2. Max Planck Institute for Infection Biology, Structural Systems Biology Group, Charitéplatz 1, 10117 Berlin, Germany;3. Heinrich Pette Institute, Leibniz Institute for Experimental Virology, Martinistraße 52, 20251 Hamburg, Germany;4. European Molecular Biology Laboratory, Hamburg Unit, Notkestraße 85, 22607 Hamburg, Germany;6. Humboldt University of Berlin, Institute for Biology-Bacterial Physiology, Philippstraße 13–Haus 22, 10115 Berlin, Germany;7. European XFEL GmbH, Sample Environment Group, Holzkoppel 4, 22869 Schenefeld, Germany;8. Faculty of Mathematics, Informatics and Natural Sciences, University of Hamburg, Rothenbaumchaussee 19, 20148 Hamburg, Germany |
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| Abstract: | Many bacteria can switch from oxygen to nitrogen oxides, such as nitrate or nitrite, as terminal electron acceptors in their respiratory chain. This process is called “denitrification” and enables biofilm formation of the opportunistic human pathogen Pseudomonas aeruginosa, making it more resilient to antibiotics and highly adaptable to different habitats. The reduction of nitrite to nitric oxide is a crucial step during denitrification. It is catalyzed by the homodimeric cytochrome cd1 nitrite reductase (NirS), which utilizes the unique isobacteriochlorin heme d1 as its reaction center. Although the reaction mechanism of nitrite reduction is well understood, far less is known about the biosynthesis of heme d1. The last step of its biosynthesis introduces a double bond in a propionate group of the tetrapyrrole to form an acrylate group. This conversion is catalyzed by the dehydrogenase NirN via a unique reaction mechanism. To get a more detailed insight into this reaction, the crystal structures of NirN with and without bound substrate have been determined. Similar to the homodimeric NirS, the monomeric NirN consists of an eight-bladed heme d1-binding β-propeller and a cytochrome c domain, but their relative orientation differs with respect to NirS. His147 coordinates heme d1 at the proximal side, whereas His323, which belongs to a flexible loop, binds at the distal position. Tyr461 and His417 are located next to the hydrogen atoms removed during dehydrogenation, suggesting an important role in catalysis. Activity assays with NirN variants revealed the essentiality of His147, His323 and Tyr461, but not of His417. |
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