Pre-steady-state kinetics of nitrogenase from Azotobacter vinelandii. Evidence for an ATP-induced conformational change of the nitrogenase complex as part of the reaction mechanism.

The Journal of Biological Chemistry
M G DuyvisH Haaker

Abstract

The pre-steady-state electron transfer reactions of nitrogenase from Azotobacter vinelandii have been studied by stopped-flow spectrophotometry. With reduced nitrogenase proteins after the initial absorbance increase at 430 nm (which is associated with electron transfer from the Fe protein to the MoFe protein and is complete in 50 ms) the absorbance decreases, which, dependent on the ratio [Av2]/[Av1], is followed by an increase of the absorbance. The mixing of reductant-free nitrogenase proteins with MgATP leads after 20 ms to a decrease of the absorbance, which could be fitted (from 0. 05 to 1 s) with a single exponential decay with a rate constant kobs = 6.6 +/- 0.8 s-1. This reaction of nitrogenase was measured at different wavelengths. The data indicate the formation of a species with a blue shift of the absorbance of metal-sulfur clusters of nitrogenase from 430 to 360 nm. The absorbance decrease at 430 nm observed (after 50 ms) in the case of the reduced nitrogenase proteins could only be simulated well if, after the initial electron transfer from the Fe protein to the MoFe protein and before dissociation of the nitrogenase complex, an additional reaction was assumed. The rate constant of this reaction was of the same or...Continue Reading

References

Dec 1, 1995·The Journal of Biological Chemistry·D Foguel, G Weber
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Jul 17, 1999·Biochimica Et Biophysica Acta·R E Sharp, S K Chapman
Sep 12, 2015·Journal of the American Chemical Society·Cedric P OwensF Akif Tezcan
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Jan 27, 2011·Biochemical Society Transactions·Thomas A ClarkeRobert R Eady

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