Localized Electronic Structure of Nitrogenase FeMoco Revealed by Selenium K-Edge High Resolution X-ray Absorption Spectroscopy

Abstract

The size and complexity of Mo-dependent nitrogenase, a multicomponent enzyme capable of reducing dinitrogen to ammonia, have made a detailed understanding of the FeMo cofactor (FeMoco) active site electronic structure an ongoing challenge. Selective substitution of sulfur by selenium in FeMoco affords a unique probe wherein local Fe–Se interactions can be directly interrogated via high-energy resolution fluorescence detected X-ray absorption spectroscopic (HERFD XAS) and extended X-ray absorption fine structure (EXAFS) studies. These studies reveal a significant asymmetry in the electronic distribution of the FeMoco, suggesting a more localized electronic structure picture than is typically assumed for iron–sulfur clusters. Supported by experimental small molecule model data in combination with time dependent density functional theory (TDDFT) calculations, the HERFD XAS data is consistent with an assignment of Fe2/Fe6 as an antiferromagnetically coupled diferric pair. HERFD XAS and EXAFS have also been applied to Se-substituted CO-inhibited MoFe protein, demonstrating the ability of these methods to reveal electronic and structural changes that occur upon substrate binding. These results emphasize the utility of Se HERFD XAS and EXAFS for selectively probing the local electronic and geometric structure of FeMoco.

Quantum Mechanics/Molecular Mechanics Study of Resting-State Vanadium Nitrogenase: Molecular and Electronic Structure of the Iron–Vanadium Cofactor

“… 35 A Mo(III) oxidation state was discovered in FeMoco via Mo X-ray absorption spectroscopy (XAS) and X-ray magnetic circular dichroism experiments and theoretical calculations. 36 − 38 A 3Fe(II)4Fe(III) oxidation state is suggested by SpReAD 33 and Se XAS 39 experiments, while theoretical calculations suggest more delocalization of electrons. 32 , 34 …”

Discussions of Fluorescence in Selenium Chemistry: Recently Reported Probes, Particles, and a Clearer Biological Knowledge

“…The data from the “edge” analysis was used to visualize selenium within the nitrogenase cofactor. This technique confirmed that the selenium was exchanged for a sulfur in FeMoco [ 146 ].…”

N2O Reductase Activity of a [Cu4S] Cluster in the 4CuI Redox State Modulated by Hydrogen Bond Donors and Proton Relays in the Secondary Coordination Sphere

“…[28] Thea dditional contribution of our system is the correlation between structure and chemical reactivity with the relevant substrate,N 2 O. Asimilar correlation between substrate activation and localization of frontier MO density has recently emerged to describe the octanuclear FeMo-cofactor of nitrogenase. [29] In our system, no visual changes are observed unless all three reaction components (1,C oCp 2 ,a nd N 2 O) are present. Because complex 1 is in the 4Cu I :1S 2À redox state,i ti su nlikely that reactivity initiates with reduction of 1 by CoCp 2 .Instead, we favor asequence where the p-accepting molecule N 2 Ob inds to 1,likely along the Cu1 À Cu2 edge,thus raising the reduction potential such that CoCp 2 can donate to the newly introduced electron holes of 1·N 2 O.…”

“…The additional contribution of our system is the correlation between structure and chemical reactivity with the relevant substrate, N 2 O. A similar correlation between substrate activation and localization of frontier MO density has recently emerged to describe the octanuclear FeMo‐cofactor of nitrogenase . In our system, no visual changes are observed unless all three reaction components ( 1 , CoCp 2 , and N 2 O) are present.…”