Duplication and extension of the Thorneley and Lowe kinetic model for Klebsiella pneumoniae nitrogenase catalysis using a mathematica software platform

Wilson, Phillip E.; Nyborg, Andrew C.; Watt, Gerald D.

doi:10.1016/s0301-4622(01)00182-x

Cited by 52 publications

(102 citation statements)

References 33 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The earlier finding that HD is formed during turnover of nitrogenase under a mixture of D 2 /N 2 , but not under D 2 alone (9, 12-16, 23), can be explained by an re mechanism in which N 2 is displaced from the E 4 (5, 6). In the absence of reactions with other substrates, this intermediate relaxes to the resting E 0 level in a two-step process: (i) release of HD to form E 2 (D) followed by (ii) release of a second HD to form E 0 .…”

Section: Resultsmentioning

confidence: 95%

“…This process releases H 2 , yielding N 2 bound to FeMocofactor that is doubly reduced relative to the resting redox level, and thereby is activated to promptly generate bound diazene (HN=NH). This mechanism predicts that during turnover under D 2 /N 2 , the reverse reaction of D 2 with the N 2 -bound product of reductive elimination would generate dideutero-E 4 2 formation, and hence a limiting stoichiometry for biological nitrogen fixation of eight electrons/protons, and provides direct experimental support for the reductive elimination mechanism.…”

mentioning

confidence: 82%

“…Nitrogenase is activated for N 2 reduction by the accumulation of four electrons/protons on its active site FeMo-cofactor, yielding a state, designated as E 4 , which contains two iron-bridging hydrides [Fe-H-Fe]. A central puzzle of nitrogenase function is an apparently obligatory formation of one H 2 per N 2 reduced, which would "waste" two reducing equivalents and four ATP.…”

mentioning

confidence: 99%

See 2 more Smart Citations

On reversible H ₂ loss upon N ₂ binding to FeMo-cofactor of nitrogenase

Yang

Khadka

Lukoyanov

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Nitrogenase is activated for N 2 reduction by the accumulation of four electrons/protons on its active site FeMo-cofactor, yielding a state, designated as E 4 , which contains two iron-bridging hydrides [Fe-H-Fe]. A central puzzle of nitrogenase function is an apparently obligatory formation of one H 2 per N 2 reduced, which would "waste" two reducing equivalents and four ATP. We recently presented a draft mechanism for nitrogenase that provides an explanation for obligatory H 2 production. In this model, H 2 is produced by reductive elimination of the two bridging hydrides of E 4 during N 2 binding. This process releases H 2 , yielding N 2 bound to FeMocofactor that is doubly reduced relative to the resting redox level, and thereby is activated to promptly generate bound diazene (HN=NH). This mechanism predicts that during turnover under D 2 /N 2 , the reverse reaction of D 2 with the N 2 -bound product of reductive elimination would generate dideutero-E 4 [E 4 to two ammonia (NH 3 ) molecules-is primarily catalyzed by the Mo-dependent nitrogenase. This enzyme comprises an electron-delivery Fe protein and MoFe protein, which contains the active site FeMo-cofactor ( Fig. 1A) (1, 2). The nitrogenase catalyzed reaction is generally thought to have the limiting stoichiometry shown in Eq. 1 (3),This equation conveys one of the most puzzling aspects of nitrogenase function, for it incorporates an obligatory formation of one mole of H 2 per mole of N 2 reduced, which requires the waste of two reducing equivalents and four ATP (1, 2). A kinetic framework for nitrogenase function that incorporates the stoichiometry of Eq. 1 is provided by the Lowe-Thorneley model (1, 2, 4), which describes transformations among catalytic intermediates (denoted E n ), where n is the number of electrons and protons (n = 0-8) delivered to MoFe protein. N 2 reduction requires activation of the MoFe protein to the E 4 state in which FeMo-cofactor has accumulated four electrons and four protons stored as two hydrides that bridge Fe atoms [Fe-H-Fe] and two protons presumably bound to sulfides of FeMo-cofactor (Fig. 1B) (5-8). The binding of N 2 to the E 4 state is coupled to the stoichiometric evolution of one H 2 per N 2 reduced.We recently presented a draft mechanism for N 2 reduction by nitrogenase that incorporates a mechanistic explanation for obligatory, reversible H 2 loss upon N 2 binding (5). We considered two models for this process, both built on our characterization of the key E 4 state, and tested them against the numerous constraints imposed by turnover under N 2 plus D 2 or T 2 (5, 9-16). In particular, these constraints include the key findings that during catalytic reduction of N 2 (see Scheme S1), a molecule of D 2 or T 2 will reduce two protons to form two HD or HT without D + /T + exchange with solvent, even though neither D 2 nor T 2 by themselves reacts with nitrogenase during turnover under Ar (5). One model, involving protonation of one of the hydrides to form H 2 , and its replacement by N 2 , was shown to violate ...

show abstract

Section: Resultsmentioning

confidence: 95%

mentioning

confidence: 82%

mentioning

confidence: 99%

See 1 more Smart Citation

On reversible H ₂ loss upon N ₂ binding to FeMo-cofactor of nitrogenase

Yang

Khadka

Lukoyanov

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

show abstract

“…Key to this issue is the finding that H 2 inhibits the reduction of diazene (22), but not hydrazine (23). We take that to mean that H 2 and diazene bind competitively, as do H 2 and N 2 (2,5,6), but that this is not the case for hydrazine. Provisionally, we further take the simplest view, that under turnover, diazene and hydrazine each join the N 2 reduction pathway at their own characteristic entry point, and then proceed to generate both H and I.…”

Section: Discussionmentioning

confidence: 99%

“…The former delivers electrons one-at-a-time to the MoFe protein, where they are utilized at the active site iron-molybdenum cofactor ([7Fe-9S-Mo-C-R-homocitrate]; FeMo-co) to reduce substrate (2,3). In the Lowe-Thorneley (LT) kinetic scheme for N 2 reduction by nitrogenase (2,5,6), the eight steps of electron/proton delivery implied by Eq. 1 are denoted E n , where n ¼ 0 to 8, with E 0 representing the resting-state enzyme.…”

Section: Epr | Eseem | Non-kramersmentioning

confidence: 99%

Unification of reaction pathway and kinetic scheme for N ₂ reduction catalyzed by nitrogenase

Lukoyanov

Yang

Barney

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

114

View full text Add to dashboard Cite

Nitrogenase catalyzes the reduction of N 2 and protons to yield two NH 3 and one H 2 . Substrate binding occurs at a complex organo-metallocluster called FeMo-cofactor (FeMo-co). Each catalytic cycle involves the sequential delivery of eight electrons/protons to this cluster, and this process has been framed within a kinetic scheme developed by Lowe and Thorneley. Rapid freezing of a modified nitrogenase under turnover conditions using diazene, methyldiazene (HN = N-CH 3 ), or hydrazine as substrate recently was shown to trap a common intermediate, designated I . It was further concluded that the two N-atoms of N 2 are hydrogenated alternately (“Alternating” ( A ) pathway). In the present work, Q-band CW EPR and 95 Mo ESEEM spectroscopy reveal such samples also contain a common intermediate with FeMo-co in an integer-spin state having a ground-state “non-Kramers” doublet. This species, designated H , has been characterized by ESEEM spectroscopy using a combination of 14,15 N isotopologs plus 1,2 H isotopologs of methyldiazene. It is concluded that: H has NH 2 bound to FeMo-co and corresponds to the penultimate intermediate of N 2 hydrogenation, the state formed after the accumulation of seven electrons/protons and the release of the first NH 3 ; I corresponds to the final intermediate in N 2 reduction, the state formed after accumulation of eight electrons/protons, with NH 3 still bound to FeMo-co prior to release and regeneration of resting-state FeMo-co. A proposed unification of the Lowe-Thorneley kinetic model with the “prompt” alternating reaction pathway represents a draft mechanism for N 2 reduction by nitrogenase.

show abstract

Biological nitrogen fixation in theory, practice, and reality: a perspective on the molybdenum nitrogenase system

Threatt

Rees

2022

FEBS Letters

View full text Add to dashboard Cite

Nitrogenase is the sole enzyme responsible for the ATP‐dependent conversion of atmospheric dinitrogen into the bioavailable form of ammonia (NH3), making this protein essential for the maintenance of the nitrogen cycle and thus life itself. Despite the widespread use of the Haber–Bosch process to industrially produce NH3, biological nitrogen fixation still accounts for half of the bioavailable nitrogen on Earth. An important feature of nitrogenase is that it operates under physiological conditions, where the equilibrium strongly favours ammonia production. This biological, multielectron reduction is a complex catalytic reaction that has perplexed scientists for decades. In this review, we explore the current understanding of the molybdenum nitrogenase system based on experimental and computational research, as well as the limitations of the crystallographic, spectroscopic, and computational techniques employed. Finally, essential outstanding questions regarding the nitrogenase system will be highlighted alongside suggestions for future experimental and computational work to elucidate this essential yet elusive process.

show abstract

Duplication and extension of the Thorneley and Lowe kinetic model for Klebsiella pneumoniae nitrogenase catalysis using a mathematica software platform

Cited by 52 publications

References 33 publications

On reversible H ₂ loss upon N ₂ binding to FeMo-cofactor of nitrogenase

On reversible H ₂ loss upon N ₂ binding to FeMo-cofactor of nitrogenase

Unification of reaction pathway and kinetic scheme for N ₂ reduction catalyzed by nitrogenase

Biological nitrogen fixation in theory, practice, and reality: a perspective on the molybdenum nitrogenase system

Contact Info

Product

Resources

About

Duplication and extension of the Thorneley and Lowe kinetic model for Klebsiella pneumoniae nitrogenase catalysis using a mathematica software platform

Cited by 52 publications

References 33 publications

On reversible H 2 loss upon N 2 binding to FeMo-cofactor of nitrogenase

On reversible H 2 loss upon N 2 binding to FeMo-cofactor of nitrogenase

Unification of reaction pathway and kinetic scheme for N 2 reduction catalyzed by nitrogenase

Biological nitrogen fixation in theory, practice, and reality: a perspective on the molybdenum nitrogenase system

Contact Info

Product

Resources

About

On reversible H ₂ loss upon N ₂ binding to FeMo-cofactor of nitrogenase

On reversible H ₂ loss upon N ₂ binding to FeMo-cofactor of nitrogenase

Unification of reaction pathway and kinetic scheme for N ₂ reduction catalyzed by nitrogenase