Spectroscopic Studies of Biological and Synthetic Tetranuclear Iron-Sulfur Clusters.

Room-temperature UV-visible adsorption, low-temperature magnetic circular dichroism (MCD), electron paramagnetic resonance (EPR), and resonance Raman (RR) spectroscopies have been used to characterize the electronic, magnetic and structural properties of the (4Fe-4S) centers in Azotobacter vinelandii Fe proteins from Mo- and V-nitrogenases (Av2 and Av2$\sp\prime$, respectively), and Bacillus subtilis glutamine phosphoribosylpyrophosphate amidotransferase. These studies have characterized the S = 1/2 and S = 3/2 forms of the (4Fe-4S) $\sp{1+}$ cluster in native Av2 and Av2$\sp\prime$ in the presence and absence of the protein unfold agent urea, glassing solvents, such as ethylene glycol and glycerol, and nucleotides, such as ATP. The results show that in the presence of glassing solvents, the (4Fe-4S) $\sp{1+}$ cluster in native Av2 is mainly in the S = 1/2 form while in the presence of urea, the cluster is mainly in the S = 3/2 form. The presence of Mg$\cdot$ATP does not effect this medium-dependent cluster spin state behavior. Av2$\sp\prime$ has a similar behavior, except that the S = 3/2 form of the cluster is more dominant in all cases. The results afford the first characterization of the MCD properties of an S = 3/2 (4Fe-4S) $\sp{1+}$ cluster and establish the changes in the electronic structure that accompany S = 3/2 $\leftrightarrow$ S = 1/2 spin state conversion. RR studies of oxidized Av2 show that the (4Fe-4S) $\sp{2+}$ is structurally analogous to similar centers in simple ferredoxins, and show that it can be partially degraded into (2Fe-2S) $\sp{2+}$ clusters, in the presence of excess thionine. Spectroscopic studies of photochemically-reduced B. subtilis amidotransferase reveal the presence of s = 3/2 (4Fe-4S) $\sp{1+}$ and S = 5/2 (4Fe-4S) $\sp{1+}$ clusters as the major and minor species, respectively, with the latter becoming more pronounced at protein high concentrations. RR studies of the native enzyme show that the structure of the (4Fe-4S) $\sp{2+}$ clusters is similar to that in simple bacterial ferredoxins. In addition, the properties of cubane-type (MoFe$\sb3$S$\sb4$) and (VFe$\sb3$S$\sb4$ synthetic clusters, which serve as possible models for the FeMo- and FeV-cofactors in Mo- and V-nitrogenase have been investigated using room-temperature UV-visible absorption, and low-temperature MCD and EPR spectroscopies. The results reveal S = 3/2 ground states for all five clusters investigated. Comparison of spectra from these synthetic clusters with the MoFe and VFe clusters at the active sites of Mo- and V-nitrogenases suggests that the differences in the spectroscopic properties of the protein bound clusters arise from differences in protein ligation