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Exploring Electron/Proton Transfer and Conformational Changes in the Nitrogenase MoFe Protein and FeMo‐cofactor Through Cryoreduction/EPR Measurements
Author(s) -
Davydov Roman,
Khadka Nimesh,
Yang ZhiYong,
Fielding Andrew J.,
Lukoyanov Dmitriy,
Dean Dennis R.,
Seefeldt Lance C.,
Hoffman Brian M.
Publication year - 2016
Publication title -
israel journal of chemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.908
H-Index - 54
eISSN - 1869-5868
pISSN - 0021-2148
DOI - 10.1002/ijch.201600026
Subject(s) - chemistry , nitrogenase , crystallography , electron paramagnetic resonance , azotobacter vinelandii , electron transfer , conformational change , cofactor , stereochemistry , nuclear magnetic resonance , photochemistry , enzyme , biochemistry , physics , organic chemistry , nitrogen fixation , nitrogen
Abstract We combine cryoreduction/annealing/EPR measurements of nitrogenase MoFe protein with results of earlier investigations to provide a detailed view of the electron/proton transfer events and conformational changes that occur during early stages of [e − /H + ] accumulation by the MoFe protein. This includes reduction of: 1) the non‐catalytic state of the iron‐molybdenum cofactor (FeMo‐co) active site that is generated by chemical oxidation of the resting‐state cofactor ( S= 3/2) within resting MoFe (E 0 ); and 2) the catalytic state that has accumulated n= 1 [e − /H + ] above the resting‐state level, denoted E 1 (1H) ( S ≥1) in the Lowe‐Thorneley kinetic scheme. FeMo‐co does not undergo a major change of conformation during reduction of oxidized FeMo‐co. In contrast, FeMo‐co undergoes substantial conformational changes during the reduction of E 0 to E 1 (1H), and of E 1 (1H) to E 2 (2H) ( S= 3/2). The experimental results further suggest that the E 1 (1H)→E 2 (2H) step involves coupled delivery of a proton and an electron (PCET) to FeMo‐co of E 1 (H) to generate a nonequilibrium S= 1 / 2 form E 2 (2H)*. This subsequently undergoes conformational relaxation and attendant change in the FeMo‐co spin state, to generate the equilibrium E 2 (2H) ( S= 3/2) state. Unexpectedly, these experiments also reveal conformational coupling between FeMo‐co and the P cluster, and between the Fe protein binding and FeMo‐co, which might play a role in gated electron transfer from reduced Fe protein to FeMo‐co.