Electron transfer stopped-flow kinetics

A comprehensive series of oxidation-reduction potential measurements have shown the FAD moiety to have the following one-electron couples PFl/PFIH = = —290 mV and PFIH 7PFIH2 = —365 mV while the FMN moiety exhibits the following PFl/PFl- = -110 mV and PFIH /PFIH = -270 mV. The FMN and FAD smiquinones were found to both be the neutral form as judged from absorption and ESR spectral data. The overlap of oxidized/semiquinone potential of the FAD moiety withkhe semiquinone/hydroquinone couple of the FMN moiety demonstrates the thermodynamic facilitation of flavin-flavin electron transfer via a one-electron mechanism. Stopped-flow kinetic data are also consistent with this view in... 

Researchers studying the stepwise kinetics of nitrogenase electron transfer using stopped-flow kinetic techniques have presented other scenarios. One hypothesis presents kinetic evidence that dissociation of Fe-protein from MoFe-protein is not necessary for re-reduction of Fe-protein by flavodoxins.13 These authors state that the possibility of ADP-ATP exchange while Fe-protein and MoFe-protein are complexed with each other cannot be excluded and that dissociation of the complex during catalysis may not be obligatory when flavodoxin is the Fe-protein reductant. This leads to the hypothesis that MgATP binds to the preformed Fe-protein/... 

M. Fabian and co-workers have studied the protein s role in internal electron transfer to the catalytic center of cytochrome c oxidase using stopped-flow kinetics. Mitochondrial cytochrome c oxidase, CcO, an enzyme that catalyzes the oxidation of ferrocytochrome c by dioxygen, is discussed more fully in Section 7.8. In the overall process, O2 is reduced to water, requiring the addition of four electrons and four protons to the enzyme s catalytic center. Electrons enter CcO from the cytosolic side, while protons enter from the matrix side of the inner mitochondrial membrane. This redox reaction. 

The midpoint redox potentials for flavin reduction in PDR are E ox/sq = nl74mV for the first couple and E sq/red = n274mV for the second couple (Gassner et al., 1995). The potential for the 2Fe-2S center is E p = nl74 mV. Thus, the thermodynamic driving force favors reduction of the 2Fe-2S center, especially by the first electron transfer. The rate of intramolecular electron transfer for the reduced flavin to the oxidized 2Fe-2S center has been estimated at >200sec based on simulation of stopped flow kinetic data (Correll et ah, 1992). 

Details of several years of UNF research exploiting rapid reaction, rapid quench and stopped flow kinetics are in the scientific literature and would be inappropriate here. The upshot was a scheme which evolved over the period and, with later refinements, is now considered to be correct in all its major features. In brief, the obligatory evolution of dihydrogen is believed to arise from the formation of a metal-dihydride complex within the reduced MoFe-protein, a two-step process requiring the transfer of two electrons successively from the MgATP-activated Fe-protein coupled with two protonation steps and the hydrolyses of four molecules of ATP to ADP N2 is bound by displacement of two... 

The stoichiometric reactions of 9 with halogens and halide ions and infrared stopped flow kinetic studies clearly show the importance of one-electron transfer in the oxidative elimination reactions between [Fe(CO)3L2] and X2 (X = Cl, Br, or I). The major steps in the mechanism are shown in... 

Knight, K. and N.S. Scrutton (2002). Stopped-flow kinetic studies of electron transfer in the reductase domain of neuronal nitric oxide synthase Reevaluation of the kinetic mechanism reveals new enzyme intermediates and variation with cytochrome P450 reductase. Biochem. J. 367,19-30. 

Stopped flow kinetic studies (30a) show that the reductase component of MMO is responsible for transferring the reducing equivalents of NADH to the hydroxylase component and that the electron-transfer steps from... 

The reactivity of metal nitrosyl complexes (51) with thiols is of particular concern in the mobilization of NO to make it accessible for the vasodilation process. Very recently, it has been reported (52) that the S-atom of cysteine reacts to bind the N-atom of the nitrosyl complex of Ru-edta to form a 1 1 intermediate species. Stopped-flow kinetic studies revealed the formation of a transient species, whose rate of formation was found to be first order with respect both [Ru (pac)(NO)] and RSH. The values of rate constants ( 1) were formd to be in the range (0.2-5) x 10 M s at 25°C. Considering the spectral features and kinetic behavior of various [Ru (pac)(SR )] and [Ru (pac)NO] species as described in the preceding sections, and analysis for the products of the above reaction (N2O), the following mechanism (Scheme 15) for the redox reactions involving electron transfer fi om thiols to coordinated NO, that results in the formation of disrdfide (RSSR) and N2O, has been proposed for the reaction of [Ru (pac)(NO)] with thiols (RSH). 

The reaction of methylcobalamin with mercury(n) is extremely rapid and has been studied by stopped flow kinetics. However, methyl transfer from methylcobalamin to tetrachloropalladate(n) is slower. In both methyl transfer reactions there are two kinetically distinct steps. The initial step involves a rapid association of the metal species with methylcobalamin, resulting in a base-on, base-off pre-equilibrium, the electrophilic metal system competing with the cobalt(m) for the lone pair of electrons on the benzimidazole nitrogen. There follows a slower demethylation step in which the electrophilic metal species attacks the Co—C bond in the uncomplexed methylcobalamin. The more rapid transfer with mercury(n) is a consequence of the greater electrophilicity of mercury(n) than palladium(ii), and the mercury(n) produces slightly more of the base-off species. ... 

Although direct complex formation is observed kinetically (stopped flow) and spectrophotometrically, where X = Br or Cl, the reaction with I results in an oxidation of the halide. The reactions are rapid and there is the question of inner- or outer-sphere electron transfer, for the [14]aneN4 complex. However, further studies (140) using ligand substituted (dimethyl) complexes reveal that for the rac-Me2[14]aneN4 isomer, two processes are observed, k = 2.9 x 104 M-1 sec-1 and a subsequent redox step, krci = 5.5 x 103 M-1 sec-1, both of which are iodide dependent. The mechanism proposed involves the formation of an octahedral complex which further reacts with a second mole of I- in the redox step ... 

Combined competition studies and stopped-flow measurements provided kinetic information on each of the two forms, L1(H20)Cr00H2+ and L Cr VOlOH2 1, in their reactions with PPh3. The hydroperoxo complex reacts by H + -catalyzed oxygen atom transfer, k 850M 2s while L1Crv(0)0H2+ reacts by electron transfer with k = AA x 105M-1 s 1. 

The results obtained in the case of primary halides were confirmed by kinetic studies of their reactions with stannylanions using a stopped flow technique. The resulting rate constants were much greater than those calculated for an electron transfer according to the Hush-Marcus theory which supports a nucleophilic reactivity rather than a single electron transfer pathway132. 

Bemasconi and co-workers have studied the kinetics of disproportion-ation-comproportionation in azaviolene systems by temperature jump, stopped-flow, and pH jump techniques, examining the electron transfer process in the light of Marcus theory.316-318... 

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