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A low-potential terminal oxidase associated with the iron-only nitrogenase from the nitrogen-fixing bacterium Azotobacter vinelandii

DOI: 10.1074/jbc.RA118.007285 DOI Help

Authors: Febin Varghese (Imperial College London) , Burak Veli Kabasakal (University of Bristol) , Charles A. R. Cotton (Max Planck Institute of Molecular Plant Physiology) , Jörg Schumacher (Imperial College London) , A. William Rutherford (Imperial College London) , Andrea Fantuzzi (Imperial College London) , James W. Murray (Imperial College London)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Journal Of Biological Chemistry

State: Published (Approved)
Published: May 2019
Diamond Proposal Number(s): 12579

Abstract: The biological route for nitrogen gas entering the biosphere is reduction to ammonia by the nitrogenase enzyme, which is inactivated by oxygen. Three types of nitrogenase exist, the least studied of which is the iron-only nitrogenase. The Anf3 protein in the bacterium Rhodobacter capsulatus is essential for diazotrophic (i.e. nitrogen-fixing) growth with the iron-only nitrogenase, but its enzymatic activity and function are unknown. Here, we biochemically and structurally characterize Anf3 from the model diazotrophic bacterium Azotobacter vinelandii. Determining the Anf3 crystal structure to atomic resolution, we observed that it is a dimeric flavocytochrome with an unusually close interaction between the heme and the flavin adenine dinucleotide cofactors. Measuring the reduction potentials by spectroelectrochemical redox titration, we observed values of -420 ± 10 mV and -330 ± 10 mV for the two FAD potentials and -340 ± 1 mV for the heme. We further show that Anf3 accepts electrons from spinach ferredoxin and that Anf3 consumes oxygen without generating superoxide or hydrogen peroxide. We predict that Anf3 protects the iron-only nitrogenase from oxygen inactivation by functioning as an oxidase in respiratory protection, with flavodoxin or ferredoxin as the physiological electron donors.

Journal Keywords: nitrogen fixation; enzyme structure; dioxygenase; oxidase; nitrogenase

Diamond Keywords: Bacteria; Enzymes

Subject Areas: Biology and Bio-materials, Chemistry

Instruments: I03-Macromolecular Crystallography

Added On: 09/05/2019 10:49

Discipline Tags:

Biochemistry Catalysis Chemistry Structural biology Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)