Abstract
Nitrogenase is the only enzyme that can cleave the triple bond in N2, making nitrogen avaiable for other organisms. It contains a complicated MoFe7S9C(homocitrate) cluster in its active site. Many computational studies with density-functional theory (DFT) of the nitrogenase enzyme have been presented, but they do not show any consensus - they do not even agree where the first four protons should be added, forming the central intermediate E4. We show that the prime reason for this is that different DFT methods give relative energies that differ by almost 600 kJ mol-1 for different protonation states. This is 4-30 times more than what is observed for other systems. The reason for this is that in some structures, the hydrogens bind to sulfide or carbide ions as protons, whereas in other structures they bind to the metals as hydride ions, changing the oxidation state of the metals, as well as the Fe-C, Fe-S and Fe-Fe distances. The energies correlate with the amount of Hartree-Fock exchange in the method, indicating a variation in the amount of static correlation in the structures. It is currently unclear which DFT method gives the best results for nitrogenase. We show that non-hybrid DFT functionals and TPSSh give the most accurate structures of the resting active site, whereas B3LYP and PBE0 give the best H2 dissociation energies. However, no DFT method indicates that a structure of E4 with two bridging hydride ions is lowest in energy, as spectroscopic experiments indicate.
Highlights
Nitrogenase (EC 1.18/19.6.1) is the only enzyme in nature that can cleave the triple bond in N2 to form ammonia and make nitrogen available for cell metabolism.[1,2,3] It is present in a few groups of bacteria and archaea.[1,2,3] The nitrogenase reaction is essential to the life on earth – 78% of the atmosphere is N2, nitrogen is often the limiting factor for plant growth and a main component of synthetic fertilisers.[3]
All structures were optimised with quantum mechanical and molecular mechanical (QM/MM) individually for each density-functional theory (DFT) method
We first compare the relative stability of the best protonation states of E4 obtained with either the B3LYP (1) or TPSS (5) functionals in our extensive study, DE15.29 As can be seen from Fig. 2, both structures are protonated on one of the m2 sulfide ions (S2B), whereas the other three protons are on the central carbide ion in 1 but on Fe ions in 5
Summary
Nitrogenase (EC 1.18/19.6.1) is the only enzyme in nature that can cleave the triple bond in N2 to form ammonia and make nitrogen available for cell metabolism.[1,2,3] It is present in a few groups of bacteria and archaea.[1,2,3] The nitrogenase reaction is essential to the life on earth – 78% of the atmosphere is N2, nitrogen is often the limiting factor for plant growth and a main component of synthetic fertilisers.[3]. Crystallographic studies have shown that the nitrogenase is a large a2b2 heterotetramer.[5,6,7,8,9] The catalytic centre is the MoFe7S9C(homocitrate) FeMo cluster bound to the protein by a cysteine and a histidine residue. The Mo ion is replaced by vanadium or iron.[10] The protein contains a Fe8S7Cys[6] cluster, called the P cluster, which transfers electrons. The electrons are provided by another protein, called the Fe protein, which binds two ATP molecules. Hydrolysis of the ATP molecules triggers the dissociation of the Fe protein, opening up for additional electron transfers
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