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Many copper containing proteins exhibit well defined ESEEM signals detected at X-band and C-band. In these systems the Cu(II) ion is coordinated to one or several histidine residues. The main sharp features measured in the ESEEM spectra originate from the interaction of the unpaired electron with the remote nitrogen nucleus of the histidine ring. The relative intensities of these features contain information about the orientation of the NQI-tensor in the molecular axis frame as defined by the principal axes of theg-matrix. This information can be related to the orientation of the imidazole ring in the complex. We present a systematic approach to determine the constraints of the Euler angles, α, β, ψ of the NQI-tensor in theg-matrix principal axis system. The first step is to analyze the intensity ratios of the quadrupole peaks and the line shape of the double quantum feature measured on the canonical positions in the EPR spectrum. This will lead to a constraint in the angles (α, β) as well as the effective hyperfine interaction. This information is further refined using spectra on other orientation selective positions. We have applied this method to Cu(II)-Bleomycin and two model compounds: Cu(II)-pypep and Cu(II)-PMA of which we have determined the principal quadrupole values and the orientation of the quadrupole tensor with respect to theg-matrix axis system.

We have investigated the action of the chemotherapeutic agent Fe(II)-bleomycin on yeast tRNA(Phe), an RNA of known three-dimensional structure. In the absence of Mg2+ ions, the RNA is cleaved preferentially at two major positions, A31 and G53, both of which are located at the terminal base pairs of hairpin loops, and coincide with the location of tight Mg2+ binding sites. A fragment of the tRNA (residues 47-76) containing the T stem-loop is also cleaved specifically at G53. Cleavage of both the intact tRNA and the tRNA fragment is abolished in the presence of physiological concentrations of Mg2+ (> 0.5 mM). Since Fe(II) is not displaced from bleomycin under these conditions, we infer that tight binding of Mg2+ to tRNA excludes productive interactions between Fe(II)-bleomycin and the RNA. These results also show that loss of cleavage is not due to Mg(2+)-dependent formation of tertiary interactions between the D and T loops. In contrast, cleavage of synthetic DNA analogs of the anticodon and T stem-loops is not detectably inhibited by Mg2+, even at concentrations as high as 50 mM. In addition, the site specificities observed in cleavage of RNA and DNA differ significantly. From these results, and from similar findings with other representative RNA molecules, we suggest that the cleavage of RNA by Fe(II)-bleomycin is unlikely to be important for its therapeutic action.

The structural and redox chemistry of the clusters [Fe4S4(SR)4]2− with R = t-alkyl have been investigated for the purpose of determining the structures of the same cluster in different environments and the stability of the oxidized species [Fe4S4(SR)4]1−. (Me3NCH2Ph)2[Fe4S4(S-t-Bu)4] crystallizes in the monoclinic space group P21/c with no imposed symmetry. (Et4N)2[Fe4S4(S-t-Bu)4] crystallizes in the tetragonal space group I 4 2m with D2d symmetry imposed on the cluster. The [Fe4S4]2+ cluster cores in both compounds exhibit compressed tetragonal structures with different extents of distortion from Td symmetry. These structures are compared to those of other [Fe4S4]2+ clusters by means of core shape parameters. Clusters with R = t-alkyl (t-Bu, C(CH3)2CH2OH, C(CH3)2CH2NHPh) in DMF exhibit, in addition to the usual 2−/3− and 3−/4− redox reactions common to all [Fe4S4(SR)4]2− species, discrete one-electron oxidations near −0.1 V vs. s.c.e. Cyclic voltammetry of [Fe4S4(S-t-Bu)4]2− reveals an essentially reversible 1−/2− couple with E 1 2 = −0.12 V, supporting the authenticity of clusters containing an oxidized ([Fe4S4]3+) core. This couple cannot be electrochemically resolved from multi-electron oxidation in the case of clusters in DMF with other types of R substituents, a behavior apparently due to cathodic potential shifts by t-alkyl groups. Stability of oxidized clusters is low, and [Fe4S4(S-t-Bu)4]1− could not be generated in appreciable concentrations at longer times by coulometric or chemical oxidation. The relative stabilities of analogue and protein [Fe4S4]3+ clusters are discussed. Preparations of four new [Fe4S4(SR)4]2− cluster salts are described including water-soluble (Et4N)2[Fe4S4(SC(CH3)2CH2OH)4].

Preparation de (Et 4 N) 3 [MoFe 3 S 4 (S-p-C 6 H 4 Cl) 3 (al 2 cat)(EtCN)](al 2 cat=diallyl-3,6catecholate). Spectre RMN de 1 H

Several properties of the FeMo-cofactor (co) of nitrogenase in N-methylformamide solution at ambient temperature have been investigated by means of 19F NMR spectroscopy. With C6H5CF3 reference signals the magnetic moment per Mo atom was found to be approximately equal to 3.9 BM, consistent with S = 3/2 ground state identified by other spectroscopic methods at low temperature. Reaction of FeMo-co with 1.0 eq of RFS- (RF = p-C6H4CF3, p-C6H4F) afforded isotropically shifted signals indicative of binding to a paramagnetic cluster. By comparison with the spectra of Fe-S and Fe-Mo-S species derivatized with RFS-, including the cubane-type MoFe3S4 clusters with S = 3/2 ground states, it was concluded that the essential FeMo-co cluster structure remains intact and a Fe atom is the probable thiolate binding site. An interaction of FeMo-co with C6H5S- had been detected earlier by low temperature EPR spectroscopy. The binding site assignment is based on large observed isotropic shifts (ca. -12ppm) compared to the much smaller values found for Mo-SRF ligands in MoFe3S4 clusters and anticipated in FeMo-co on the basis of recent spectroscopic results. Isotropic 19F shifts have proven extremely sensitive to electronic and structural features of Fe-S and Fe-Mo-S clusters. The inclusion of a 19F NMR label in FeMo-co should prove of utility in further investigation of cofactor properties and reactions.