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Reduction of N2 by supported tungsten clusters gives a model of the process by nitrogenase.

ABSTRACT: Metalloenzymes catalyze difficult chemical reactions under mild conditions. Mimicking their functions is a challenging task and it has been investigated using homogeneous systems containing metal complexes. The nitrogenase that converts N(2) to NH(3) under mild conditions is one of such enzymes. Efforts to realize the biological function have continued for more than four decades, which has resulted in several reports of reduction of N(2), ligated to metal complexes in solutions, to NH(3) by protonation under mild conditions. Here, we show that seemingly distinct supported small tungsten clusters in a dry environment reduce N(2) under mild conditions like the nitrogenase. N(2) is reduced to NH(3) via N(2)H(4) by addition of neutral H atoms, which agrees with the mechanism recently proposed for the N(2) reduction on the active site of nitrogenase. The process on the supported clusters gives a model of the biological N(2) reduction.

SUBMITTER: Murakami J

PROVIDER: S-EPMC3350986 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Reduction of N2 by supported tungsten clusters gives a model of the process by nitrogenase.

Murakami Junichi J Yamaguchi Wataru W

Scientific reports 20120514

Metalloenzymes catalyze difficult chemical reactions under mild conditions. Mimicking their functions is a challenging task and it has been investigated using homogeneous systems containing metal complexes. The nitrogenase that converts N(2) to NH(3) under mild conditions is one of such enzymes. Efforts to realize the biological function have continued for more than four decades, which has resulted in several reports of reduction of N(2), ligated to metal complexes in solutions, to NH(3) by prot ...[more]

PMID: 22586517

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Project description:Nitrogenase reduces dinitrogen (N2) by six electrons and six protons at an active-site metallocluster called FeMo cofactor, to yield two ammonia molecules. Insights into the mechanism of substrate reduction by nitrogenase have come from recent successes in trapping and characterizing intermediates generated during the reduction of protons as well as nitrogenous and alkyne substrates by MoFe proteins with amino acid substitutions. Here, we describe an intermediate generated at a high concentration during reduction of the natural nitrogenase substrate, N2, by wild-type MoFe protein, providing evidence that it contains N2 bound to the active-site FeMo cofactor. When MoFe protein was frozen at 77 K during steady-state turnover with N2, the S = 3/2 EPR signal (g = [4.3, 3.64, 2.00]) arising from the resting state of FeMo cofactor was observed to convert to a rhombic, S = 1/2, signal (g = [2.08, 1.99, 1.97]). The intensity of the N2-dependent EPR signal increased with increasing N2 partial pressure, reaching a maximum intensity of approximately 20% of that of the original FeMo cofactor signal at > or = 0.2 atm N2. An almost complete loss of resting FeMo cofactor signal in this sample implies that the remainder of the enzyme has been reduced to an EPR-silent intermediate state. The N2-dependent EPR signal intensity also varied with the ratio of Fe protein to MoFe protein (electron flux through nitrogenase), with the maximum signal intensity observed with a ratio of 2:1 (1:1 Fe protein:FeMo cofactor) or higher. The pH optimum for the signal was 7.1. The N2-dependent EPR signal intensity exhibited a linear dependence on the square root of the EPR microwave power in contrast to the nonlinear response of signal intensity observed for hydrazine-, diazene-, and methyldiazene-trapped states. 15N ENDOR spectroscopic analysis of MoFe protein captured during turnover with 15N2 revealed a 15N nuclear spin coupled to the FeMo cofactor with a hyperfine tensor A = [0.9, 1.4, 0.45] MHz establishing that an N2-derived species was trapped on the FeMo cofactor. The observation of a single type of 15N-coupled nucleus from the field dependence, along with the absence of an associated exchangeable 1H ENDOR signal, is consistent with an N2 molecule bound end-on to the FeMo cofactor.

Project description:Nitrogenase is a fascinating enzyme in biology that reduces dinitrogen from air to ammonia through stepwise reduction and protonation. Despite it being studied in detail by experimental and computational groups, there are still many unknown factors in the catalytic cycle of nitrogenase, especially related to the addition of protons and electrons and their order. A recent biomimetic study characterized a potential dinitrogen-bridged diiron cluster as a synthetic model of nitrogenase. Using strong acid and reductants, the dinitrogen was converted into ammonia molecules, but details of the mechanism remains unknown. In particular, it was unclear from the experimental studies whether the proton and electron transfer steps are sequential or alternating. Moreover, the work failed to establish what the function of the diiron core is and whether it split into mononuclear iron fragments during the reaction. To understand the structure and reactivity of the biomimetic dinitrogen-bridged diiron complex [(P2P'PhFeH)2(μ-N2)] with triphenylphosphine ligands, we performed a density functional theory study. Our computational methods were validated against experimental crystal structure coordinates, Mössbauer parameters, and vibrational frequencies and show excellent agreement. Subsequently, we investigated the alternating and consecutive addition of electrons and protons to the system. The calculations identify a number of possible reaction channels, namely, same-site protonation, alternating protonation, and complex dissociation into mononuclear iron centers. The calculations show that the overall mechanism is not a pure sequential set of electron and proton transfers but a mixture of alternating and consecutive steps. In particular, the first reaction steps will start with double proton transfer followed by an electron transfer, while thereafter, there is another proton transfer and a second electron transfer to give a complex whereby ammonia can split off with a low energetic barrier. The second channel starts with alternating protonation of the two nitrogen atoms, whereafter the initial double proton transfer, electrons and protons are added sequentially to form a hydrazine-bound complex. The latter split off ammonia spontaneously after further protonation. The various reaction channels are analyzed with valence bond and orbital diagrams. We anticipate the nitrogenase enzyme to operate with mixed alternating and consecutive protonation and electron transfer steps.

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Reduction of N2 by supported tungsten clusters gives a model of the process by nitrogenase.

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Reduction of N2 by supported tungsten clusters gives a model of the process by nitrogenase.

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