EPR-Silent States

The intermediate states Ni-SU, Ni-Slr, Ni-Sla, and Ni-R, where the number of eleetrons in the [NiFe] eenter is even, are commonly denoted as EPR-silent states. X-ray absorption speetroscopy (XAS) experiments are compatible with a formal Ni in the EPR-silent states (Ni-SU Ni-Sl Ni-R) [38,41]. WithNi L-edge XAS it was found that the Ni is most likely in a high-spin state (5 = 1) [95]. This has so far not been eorroborated by EPR spectroscopy, possibly because of the presenee of a large zero-field splitting that makes it impossible to detect the signal at X-band frequeneies (9 GHz). 

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In the presence of redox mediators, the Nia-C state can be further reduced to an EPR-silent state (by increasing the H2 partial pressure) in a reaction requiring one electron and one proton. Assuming trivalent nickel in the Nig-C " state, this reaction can be most simply interpreted as... 

Methyl coenzyme M reductase has been extracted from strains of the thermophilic bacterium M. thermoautotrophicum and intensively studied. When isolated, it contains both CoM and CoB [111,112], The enzyme is irreversibly inactivated by oxygen. Even when extracted anaerobically it loses activity within a few hours [113], producing the inactive, EPR-silent state MCRSI C1 1 in which the nickel is presumably Ni(II). 

For [NiFe] hydrogenase, many mechanisms of the catalytic reaction have been suggested. 36,37 One of these schemes is shown in Figure 9-3.37,38 The oxidized system (Ni-A and Ni-B) is reduced under an atmosphere of H2 and switches to a reduced system. This scheme has four paramagnetic states Ni-A, Ni-B, Ni-C and Ni-L and three EPR-silent states Ni-SU, Ni-SI and Ni-R. Volbeda et al. have found that Ni-SI has the two different states, denoted by Ni-SI and Ni-SIn.10,11 The Ni-A... 

Ni-R is also an EPR-silent state and has the redox state Ni(II)Fe(II). Hall et al. have shown that Ni-R has H2 bound to the Fe atom by DFT calculations and considering the electronic states.51,62-64 Gioia et al. and Stein et al. propose similar structures.66,67 Both groups state that the bridging ligand X is a H atom and Ni-R... 

Figure 9-5(a). Structure of the EPR-silent states, such as Ni-SU, Ni-SI, and Ni-R of Pavlov et al. and Hall et al. from theoretical investigations. Note that Hall et al. suggest the existence of two Ni-SI states, Ni-SIa and Ni-SIb... 

In the case of the [FeNij-hydrogenase of Desulfovibrio gigas IR absorption in the 1900-2100 cm region has been assigned for each of the redox-active states [44], These data are collected in Table 2. This study revealed the presence of an EPR silent state labeled SU that exists as a mixture of two protonation states. The changes in IR properties have been used to determine the electrochemical potentials required for the changes in redox state. These data are presented in the following section. 

In the dye-oxidized EPR-silent state a cluster that was diamagnetic in the dithionite-reduced state became paramagnetic, with MCD and magnetization characteristics assignable to an S = 5/2 spin system with near axial symmetry. These data are very similar, but are not identical with those for the P clusters of MoFe proteins and suggest the presence of analogous clusters in the VFe proteins. 

The formation of mixed-valence intermediates is not limited to purely electron-transfer redox series, as pointed out before and as described below however, the overall reversibility is typically checked by regenerating the starting spectrum to 100% after completing the backreaction. If the method is selective only for the intermediate state, as, e.g., for EPR active odd-electron species in equilibrium with EPR-silent states, or if the two-step redox system can be analysed via stepwise monitoring, " the spectral features of an intermediate as distinct from those of the neighbouring redox states can be well identified. A case in point is discussed further below by example of intermediates [(C R )M(p-L)M(C R )]". ... 

FIGURE 7.5 Schematic view of the [Ni-Fe] hydrogenase catalytic cycle, as suggested in Ref. [2]. The EPR detectable states are shown in italics the EPR silent states are shown with bold fonts. The exact mechanism and the transient species involved are not yet known. More details in the text. 

When the enzyme is turning over the EPR signal essentially disappears, leaving an EPR-silent state in which the FeMoco site is super-reduced to what is presumed to be its catalytically active form. In addition, a third state in which the 5 = 1 EPR signal disappears is produced upon oxidation under non-turnover conditions. Thus the M center within the protein shows three states of oxidation, and these appear to have been reproduced in the FeMoco extracted from the protein ... 

A similar model, again using a terminal SMe to represent the cysteine that connects the 2Fe unit to the 4Fe4S cluster, was employed by Liu and Hu in their attempts to characterize the possible structures of the active site of [Fe]H2ase at different redox levels. According to their study, the fully oxidized, inactive, and EPR-silent state of the enzyme was satisfied by an Fe pe model with a terminally bound OH group on the distal Fe. The oxidized active state of the... 

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