Metal-Bound Histidine Modes in UV Resonance Raman Spectra of Cu, Zn Superoxide Dismutase

Abstract

Ultraviolet resonance Raman [UVRR] spectra of Cu, Zn superoxide dismutase [SOD] contain bands arising from vibrations of metal-bound histidine ligands. Spectra in H 2 O solution reveal several modes of the His61 side chain, which bridges the Cu 2+ and Zn 2+ ions as imidazolate. The disappearance of these bands signals disruption of the bridge when the pH is lowered to 3.0, or the Cu 2+ is reduced to Cu + . Binding of hydroxide [pH 12] or cyanide to the Cu 2+ perturbs the imidazolate modes, reflecting geometry changes induced by these strong-field ligands. In D 2 O solution several additional bands become enhanced which arise from histidine ligands that have undergone NH/D exchange. Some of these are attributed to Cu-bound ligands and others to Zn-bound ligands, on the basis of selective changes accompanying removal and replacement of the metals. Excitation profiles are similar for these bands, and for the bridging imidazolate bands; they are redshifted relative to nonligating histidine. The detection of site-specific histidine ligand modes gives promise for wide applicability of UVRR spectroscopy in studying histidine ligation in metalloproteins. The single tyrosine residue of SOD, which is a target of active-site-catalyzed nitration by peroxynitrite, is found to have an elevated pK a , 11.4, despite being exposed to solvent.

Raman Spectroscopy of Proteins

“…The band at 282 cm 1 in the UVRR spectra of Cd(II)-bound zinc-finger peptides has been suggested to be the metal-ligand stretching mode. 58 59,60 The UVRR bands of amino acid residues described here, which would serve as useful structural markers, are also summarized in Table 2 together with their vibrational characters.…”

New Insights into the Coordination Chemistry and Molecular Structure of Copper(II) Histidine Complexes in Aqueous Solutions

“…To elucidate the molecular structures of Cu 2+ /his complexes and to obtain insight into the forces that drive anchoring of the metal ion, many studies have been carried out since the late 1960s. In these studies, a wide variety of techniques have been used to obtain the desired information, ranging from spectroscopic techniques, like circular dichroism, 10,25,26 ultraviolet/visible/ near-infrared (UV/vis/NIR), 10,25-29 infrared (IR), 3,14,[26][27][28][29][30][31][32][33] Raman, [3][4][5]10,13,15-17,33-35 1 H NMR, 10,32,36-39 13 C NMR, 36 electron spin resonance (ESR), 10,27,37,40-48 and extended X-ray absorption fine structure (EXAFS), 6,49 to potentiometric 10,26,28,50-54 and calorimetric 55 experiments. Several Cu 2+ /his complexes consisting of one or two central Cu atoms have been proposed in these studies, with one or two his ligands in a mono-, bi-, or tridentate coordination and neutrally, negatively, or positively charged.…”

Raman structural markers of tryptophan and histidine side chains in proteins

“…Although a characteristic Raman band is seen at 1440 -1434 cm Ϫ1 for the [N -M, N -H] form, 68 it will be overlapped by COH bending bands of aliphatic side chains in visible Raman spectra 76 -78 and by X-Pro imide II bands in the UV Raman spectra of proteins that contain Pro residues. 69,71 The utility of the 1440 -1434 cm for N -metal-bound His residues in proteins: two His residues of a Zn finger 74 and the Febound proximal His residues of hemoglobin and microperoxidases. 72 However, UV Raman bands have also been found in the 1365-1344 cm Ϫ1 region for plastocyanin, 61 of which two His residues are bound to a Cu ion via N but not via N , and for Cu-depleted SOD, 71 in which the remaining Zn ion is expected to bind to the N atoms of three His residues in the active site.…”

“…69,71 The utility of the 1440 -1434 cm for N -metal-bound His residues in proteins: two His residues of a Zn finger 74 and the Febound proximal His residues of hemoglobin and microperoxidases. 72 However, UV Raman bands have also been found in the 1365-1344 cm Ϫ1 region for plastocyanin, 61 of which two His residues are bound to a Cu ion via N but not via N , and for Cu-depleted SOD, 71 in which the remaining Zn ion is expected to bind to the N atoms of three His residues in the active site. The imidazolate bridge ([N -M, N -MЈ]) is characterized by a strong Raman band that is attributable to a stretch mode of the C 2 ON OC 4 linkage 69,81 at 1292-1282 cm Ϫ1 in both visible and UV Raman spectra.…”