Investigation of complex formation and the FeV cofactor of Vanadium Nitrogenase

Abstract: Life on Earth depends on the availability of fixed nitrogen, which is an essential building block for – among others – DNA and proteins. It is the limiting factor for biomass production. Nitrogenase is the only enzyme that is capable of biological nitrogen fixation. Despite decades of research, its reduction mechanism remains enigmatic. The Haber-Bosch process, developed in the early 20th century, is the industrial way of nitrogen fixation and is used to produce fertilizers. Currently it provides approximately 50% of all fixed nitrogen on Earth. It enabled the steep growth in human population and strongly decreased the prevalence of famines. But it comes at the cost of environmental pollution through the input of reactive nitrogen species to our soils by surplus fertilizers and a high energy consumption that poses a non-negligible contribution to climate change.
A detailed understanding of the mechanistic principles that underlie nitrogenase catalysis as well as a profound knowledge of the factors that stir differences in reactivity between the three known nitrogenases can guide the development of biomimetic catalysts or biotechnological applications of the nitrogenase system to provide an environmentally friendly source of fixed nitrogen. Harnessing the potential of the V nitrogenase system to reduce CO to hydrocarbons would pose an additional interesting application.
To contribute to the field, this work investigated the active site of FeV cofactor utilizing mutational studies, activity assays, turnover assays and X-ray crystallography. Mutation of the active site residues Q176D and H180D provided insights into their mechanistic function and pointed towards homocitrate as an additional proton supplier during catalysis. Possible differences in the reduction of acetylene compared to CO or N2 were highlighted. To this point, the proposed CO reduction mechanism of V nitrogenase is consistent with the obtained experimental results. Due to a severe instability of mutated proteins, crystallographic analysis of crystals obtained after turnover was not successful in all cases. The individual oxidation states of the seven Fe ions in FeV cofactor were determined with spatially resolved anomalous dispersion refinement and compared to FeMo cofactor. Complex formation of VFe protein with all three Fe proteins was investigated by cryo-electron microscopy. The structural analysis showed a high similarity to complex formation in Mo nitrogenase. A possible role for the additional VnfG subunit as a mediator in cross-compatibility of the catalytic component with different Fe proteins is proposed

Angaben zum Objekt

• Standort: Deutsche Nationalbibliothek Frankfurt am Main
• Umfang: Online-Ressource
• Ausgabe: 1, Version
• Sprache: Englisch
• Anmerkungen: Universität Freiburg, Dissertation, 2023

Klassifikation und Themen

• Schlagwort: Stickstofffixierung ; Azotobacter vinelandii ; Nitrogenase ; Vanadium ; Kristallographie ; Kryoelektronenmikroskopie ; Strukturbiologie ; Molekularbiologie ; Stickstofffixierung ; Mikrobiologie ; Strukturaufklärung ; Metallcluster

Beteiligte, Orts- und Zeitangaben

• Ereignis: Veröffentlichung
• (wo): Freiburg
• (wer): Universität
• (wann): 2023
• Urheber: Parison, Katharina
• Beteiligte Personen und Organisationen: Einsle, Oliver ; Einsle, Oliver ; Andrade, Susana ; Albert-Ludwigs-Universität Freiburg. Institut für Biochemie ; Albert-Ludwigs-Universität Freiburg. Fakultät für Chemie und Pharmazie

Weitere Informationen

• DOI: 10.6094/UNIFR/242201
• URN: urn:nbn:de:bsz:25-freidok-2422016
• Rechteinformation: Open Access; Der Zugriff auf das Objekt ist unbeschränkt möglich.
• Letzte Aktualisierung: 25.03.2025, 13:54 MEZ