Influence of pN2 and pD2 on HD formation by various nitrogenases

Li, Jilun; Burris, R. H.

doi:10.1021/bi00288a019

Cited by 54 publications

(74 citation statements)

References 16 publications

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“…As indicated in Fig. 1 and in Discussion, reduction of C 2 H 2 by E 4 (2D) is predicted to yield C 2 H 2 D 2 and possibly C 2 H 3 D, whereas reduction by E 2 (D) would yield C 2 H 3 D. These studies used 13 C 2 H 2 to avoid confusion from the small but significant natural abundance of 13 C (1.07%) (25) in acetylene.…”

Section: Resultsmentioning

confidence: 94%

“…2), equivalent to 2% of the amount of 13 C 2 H 4 generated. Most significantly, a product with m/z of 32, corresponding to 13 was cleanly detected (Fig. 2, Inset), with a lower, although still significant, production rate of ∼1.9 nmol 13 C 2 H 2 D 2 /nmol MoFe protein over 20 min.…”

Section: Resultsmentioning

confidence: 97%

“…2, Inset), with a lower, although still significant, production rate of ∼1.9 nmol 13 C 2 H 2 D 2 /nmol MoFe protein over 20 min. This species can only have been produced by 13 C 2 H 2 intercepting E 4 (2D) (Fig. 1), the intermediate formed through N 2 replacement by D 2 .…”

Section: Resultsmentioning

confidence: 99%

“…Ultrapure helium was purchased from Airgas. Ethylene (99.9% vol/ vol) was obtained from Praxair Inc. 13 C-acetylene (99 atom % 13 C) and D 2 (99.8 atom % D) were purchased from Sigma-Aldrich and Isotec, respectively. All other reagents were obtained from Sigma-Aldrich or Fisher Scientific and were used without further purification, unless specified otherwise.…”

Section: Methodsmentioning

confidence: 99%

“…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)(10)(11)(12)(13)(14)(15)(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).…”

mentioning

confidence: 99%

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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.

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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 ...

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Section: Resultsmentioning

confidence: 94%

Section: Resultsmentioning

confidence: 97%

Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

See 3 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.

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Legume Nitrogen Fixation and Soil Abiotic Stress: From Physiology to Genomics and Beyond

Valentine

Benedito

Kang

2018

Annual Plant Reviews Online

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Legumes are important components of the nitrogen cycle on land. Agricultural systems have traditionally relied much on legumes for nitrogen input because many species are able to establish symbioses with diazotrophic bacteria (rhizobia) and thus trade metabolites and reduced compounds. Photosynthates produced in the leaves are allocated to the root nodule to supply the bacteroids with carbon, in exchange for reduced nitrogen (ammonia) produced by the rhizobia from atmospheric nitrogen. Despite its major significance to plant breeding and sustainable agriculture, the impact of abiotic stresses on nodule development and stability and on symbiotic nitrogen fixation remains poorly understood, particularly at the molecular level. However, the study of model legume species and the development of a plethora of resources, particularly the elucidation of the genome sequences of three legume species, are now revealing many traits of agricultural importance in legumes as well as other aspects that are not easily studied in other plant models, such asArabidopsisor rice. In this chapter, we will discuss the effects of abiotic stresses, such as drought, phosphate deficiency and aluminium toxicity, on symbiotic nitrogen fixation and provide perspectives on molecular approaches to the analysis of stress responses in legumes.

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