Proton NMR study of structural perturbation in sperm whale myoglobin due to pentaammineruthenium(III) groups appended to surface histidyl imidazoles

Abstract

[a5Ru]3Mb (a = NH3), which exhibits novel catalytic properties, and its model compound [a5RuImH] 3•2 20 (ImH = imidazole) have been investigated by 360-MHz *H NMR. The model compound in 2H20 exhibits three nonlabile single-proton peaks corresponding to 2'-H, 5'-H, and 4'-H of the coordinated imidazole at -31.2, -2.5, and near 4.7 ppm at 25 °C and pH 6.1, respectively. A pH titration gives a single pK 8.4 for deprotonation of the imidazole, with resulting increases of imidazole contact shifts. Ruthenium-labeled Mb and Ru-labeled apo-Mb also exhibit peaks at -36 ppm, which are assigned to the 2'-H of the imidazoles of His-12, His-81, and His-113. Integration of these 2'-H peaks relative to a heme methyl allows determination of the extent of reaction of surface histidines with pentaammineruthenium(III). Detailed comparison of the hyperfme shifted resonances for metaquo, methydroxy, metcyano, metazide, and deoxy forms of the proteins show negligible influence of the Ru chromophores on shifts, indicating protein folding essentially unaltered from that in the native protein. The difference in the high-spin/low-spin separation in the metazide form for the native and Ru-labeled proteins is estimated at only 24 cal, indicating a slightly weaker axial ligand field for the latter derivative. The decreased proton spin-lattice relaxation times and increased line widths of Ru-labeled Mb relative to native Mb complexes indicate the presence of metal-metal interactions which influence the electron-spin relaxation of the iron center.

Proton NMR characterization of Chromatium gracile high-potential iron protein and its ruthenium-modified derivatives. Modulation of the reduction potentials in low- and high-potential [Fe4S4] ferredoxins

“…The absence of significant perturbations in the NMR spectrum of a protein following ruthenium modification has been noted previously for myoglobin (Toi et al, 1984). In that case, no chemical shift changes were observed, although slight enhancements of the relaxation rates were attributed to a minor interaction between the two metal centers.…”

Nuclear magnetic resonance of paramagnetic metalloproteins

“…738 The heme shifts remain unaltered upon ruthenium binding. Both a line broadening and a shortening of the TI values of the heme signals are noticed in the ruthenium derivatives, these being sizable in the lowspin cases.…”

1H NMR study of the solvolysis of the paramagnetic tetrachloro-bis(imidazole)ruthenium(III) anion in water, methanol, and dimethyl sulfoxide

“…When this work was undertaken, 'H NMR spectra had been reported for aqueous solutions of [Ru(NH,),Im]Cl,, used as a model for ruthenium-histidyl protein complexes (13). More recently, the spectra of (ImH) [RuC1,1m2] in DMSO (10) and of Na[RuCl,(DMSO)Im] in D,O (14) were reported.…”