scholarly journals Nitrogenase from Klebsiella pneumoniae. An e.p.r. signal observed during enzyme turnover under ethylene is associated with the iron-molybdenum cofactor

1983 ◽  
Vol 211 (2) ◽  
pp. 495-497 ◽  
Author(s):  
T R Hawkes ◽  
D J Lowe ◽  
B E Smith
Keyword(s):  
Electron Paramagnetic Resonance ◽  
Klebsiella Pneumoniae ◽  
Isotopic Substitution ◽  
Resonance Signal ◽  
Paramagnetic Resonance ◽  
Mofe Protein ◽  
Femo Cofactor ◽  
Enzyme Turnover ◽  
Iron Molybdenum Cofactor ◽  
Electron Paramagnetic

During turnover at 10 degrees C at pH 7.4 in the presence of ethylene, the MoFe protein of Klebsiella pneumoniae nitrogenase (Kp 1) exhibited an electron-paramagnetic-resonance signal with g-values at 2.12, 1.998 and 1.987. 57Fe isotopic substitution demonstrated that this signal arose from the Kp 1 FeMo-cofactor in an S = 1/2 spin state.

scholarly journals Evidence on intramolecular electron transfer in the MoFe protein of nitrogenase from Klebsiella pneumoniae from rapid-freeze electron-paramagnetic-resonance studies of its oxidation by ferricyanide

1983 ◽  
Vol 209 (1) ◽  
pp. 207-213 ◽  
Author(s):  
B E Smith ◽  
D J Lowe ◽  
G X Chen ◽  
M J O'Donnell ◽  
T R Hawkes
Keyword(s):  
Electron Transfer ◽  
Klebsiella Pneumoniae ◽  
Similar Rate ◽  
Intramolecular Electron Transfer ◽  
Paramagnetic Resonance ◽  
Oxidation Level ◽  
Mofe Protein ◽  
Enzyme Turnover ◽  
Electron Paramagnetic ◽  
Intramolecular Process

A transient e.p.r. signal with g-values of 2.05, 1.95 and 1.81 was observed in rapid-freezing experiments when Kpl, the MoFe protein of nitrogenase from Klebsiella pneumoniae, was oxidized by ferricyanide or by some dyes. This e.p.r. signal was assigned to the ‘P’-centres and, since such signals are characteristics of [4Fe-4S]1+ clusters, provides further evidence for the ‘P’-centres being in the [4Fe-4S]0 oxidation level in the dithionite-reduced protein. When 4-10-fold excesses of ferricyanide were used as oxidants, the rate of disappearance of the transient e.p.r. signal was independent of the concentrations of ferricyanide, Kpl or ferrocyanide, i.e. its disappearance was by an intramolecular process. Under some circumstances the g = 3.7 e.p.r. signal from the FeMo-cofactors disappeared at a similar rate. It was concluded that, in these circumstances, the g = 3.7 e.p.r. signal disappears, owing to intramolecular electron transfer to the ‘P’-centres in the [4Fe-4S]2+ (P2+) oxidation level, whereas the gav. = 1.933 transient e.p.r. signal from the P1+ centres disappears, owing to a change in its spin state from S = 1/2 to S = 5/2 the rate of this process being maximal when there are two P1+ centres in the half-molecule. The rate of the intramolecular decay of the e.p.r. signals, 4.1 +/- 0.8 s-1, is the same as the rate of enzyme turnover. It is suggested that both processes may be linked to the same conformational change, which triggers, or is triggered by, intramolecular electron transfer.

scholarly journals Electron-paramagnetic-resonance studies on nitrogenase of Klebsiella pneumoniae. Evidence for acetylene- and ethylene-nitrogenase transient complexes

1978 ◽  
Vol 173 (1) ◽  
pp. 277-290 ◽  
Author(s):  
D J Lowe ◽  
R R Eady ◽  
R N F Thorneley
Keyword(s):  
Electron Paramagnetic Resonance ◽  
Klebsiella Pneumoniae ◽  
Binding Site ◽  
Dissociation Constants ◽  
Paramagnetic Resonance ◽  
Fe Protein ◽  
Low Concentrations ◽  
Direct Binding ◽  
Electron Paramagnetic ◽  
Single Binding Site

Klebsiella pneumoniae nitrogenase exhibited four new electron-paramagnetic-resonance signals during turnover at 10 degrees C, pH7.4, which were assigned to intermediates present in low concentrations in the steady state. 57Fe-substituted Mo–Fe protein showed that they arose from Fe–S clusters in the Mo–Fe protein of nitrogenase. The new signals are designated: Ic, g values at 4.67, 3.37 and approx. 2.0; VI, g values at 2.125, 2.000 and 2.000; VII, g values at 5.7 and 5.4; VIII, g values at 2.092, 1.974 and 1.933. The sharp axial signal VI arises from a Fe4S4 cluster at the −1 oxidation level. This signal was only detected in the presence of ethylene and provides the first evidence of an enzyme–product complex for nitrogenase. [13C]Acetylene and [13C]ethylene provided no evidence for direct binding of this substrate and product to the Fe–S clusters giving rise to these signals. The dependence of signal intensities on acetylene concentration indicated two types of binding site, with apparent dissociation constants K less than 16 micron and K approximately 13mM. A single binding site for ethylene (K=1.5mM) was detected. A scheme is proposed for the mechanism of reduction of acetylene to ethylene and inhibition of this reaction by CO.

Reconstitution of the 2Fe-2S Center andg= 1.89 Electron Paramagnetic Resonance Signal into OverproducedNostocsp. PCC 7906 Rieske Protein†

Biochemistry ◽  
1996 ◽  
Vol 35 (48) ◽  
pp. 15485-15493 ◽  
Author(s):  
Beatrice Holton ◽  
Xiaonan Wu ◽  
Alexandre I. Tsapin ◽  
David M. Kramer ◽  
Richard Malkin ◽  
...  
Keyword(s):  
Electron Paramagnetic Resonance ◽  
Electron Paramagnetic Resonance Signal ◽  
Resonance Signal ◽  
Rieske Protein ◽  
Paramagnetic Resonance ◽  
Electron Paramagnetic
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