Connecting nitrogenase intermediates with the kinetic scheme for N2 reduction by a relaxation protocol and identification of the N2 binding state.

Connecting nitrogenase intermediates with the kinetic scheme for N2 reduction by a relaxation protocol and identification of the N2 binding state. Research Abstract Details 

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Connecting nitrogenase intermediates with the kinetic scheme for N2 reduction by a relaxation protocol and identification of the N2 binding state. Abstract Text:

Dmitriy Lukoyanov,Brett M Barney,Dennis R Dean,Lance C Seefeldt,Brian M Hoffman,

A major obstacle to understanding the reduction of N2 to NH3 by nitrogenase has been the impossibility of synchronizing electron delivery to the MoFe protein for generation of specific enzymatic intermediates. When an intermediate is trapped without synchronous electron delivery, the number of electrons, n, it has accumulated is unknown. Consequently, the intermediate is untethered from kinetic schemes for reduction, which are indexed by n. We show that a trapped intermediate itself provides a "synchronously prepared" initial state, and its relaxation to the resting state at 253 K, conditions that prevent electron delivery to MoFe protein, can be analyzed to reveal n and the nature of the relaxation reactions. The approach is applied to the "H+/H- intermediate" (A) that appears during turnover both in the presence and absence of N2 substrate. A exhibits an S = 1/2 EPR signal from the active-site iron-molybdenum cofactor (FeMo-co) to which are bound at least two hydrides/protons. A undergoes two-step relaxation to the resting state (C): A --> B --> C, where B has an S = 3/2 FeMo-co. Both steps show large solvent kinetic isotope effects: KIE approximately 3-4 (85% D2O). In the context of the Lowe-Thorneley kinetic scheme for N2 reduction, these results provide powerful evidence that H2 is formed in both relaxation steps, that A is the catalytically central state that is activated for N2 binding by the accumulation of n = 4 electrons, and that B has accumulated n = 2 electrons.

Connecting nitrogenase intermediates with the kinetic scheme for N2 reduction by a relaxation protocol and identification of the N2 binding state. Publishing Authors By Initials

D Lukoyanov,BM Barney,DR Dean,LC Seefeldt,BM Hoffman,

For similar biochemical phenomena, metabolism, and nutrition: biochemical phenomena: substrate specificity research abstracts see: biochemical phenomena, metabolism, and nutrition: biochemical phenomena: substrate specificity research

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Connecting nitrogenase intermediates with the kinetic scheme for N2 reduction by a relaxation protocol and identification of the N2 binding state. Journal Published:

PUBLICATION TYPE: Research Support, U.S. Gov't,

Journal: Proceedings of the National Academy of Sciences of

VOLUME: 104

Page Numbers: 1451-5

Journal Abbreviation: Proc. Natl. Acad. Sci. U.S.A.

ISSN: 0027-8424

DAY: 24

MONTH: 01

YEAR: 2007

Connecting nitrogenase intermediates with the kinetic scheme for N2 reduction by a relaxation protocol and identification of the N2 binding state. Information

Number of References:

LANGUAGE: eng

NlmUniqueID: 7505876

Connecting nitrogenase intermediates with the kinetic scheme for N2 reduction by a relaxation protocol and identification of the N2 binding state. Keywords Mesh Terms:

KEYWORDS: Substrate Specificity

MESH TERMS: chemistry

Chemical & Substance for Abstract: Connecting nitrogenase intermediates with the kinetic scheme for N2 reduction by a relaxation protocol and identification of the N2 binding state. Information

Substance Name: Nitrogenase

Registry Number: EC 1.18.6.1

Grant and Affiliation Information for Connecting nitrogenase intermediates with the kinetic scheme for N2 reduction by a relaxation protocol and identification of the N2 binding state.

AFFILIATION: Department of Chemistry, Northwestern University, Evanston, IL 60208, USA.

Country: United States

AGENCY: United States NIGMS

GRANT: R01-GM59087

ACRONYM: GM

MEDLINETA: Proc Natl Acad Sci U S A

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