Four- and five-coordinate species in nickel-reconstituted hemoglobin and myoglobin: Raman identification of the nickel-histidine stretching mode

Abstract

Nickel(II)-reconstituted hemoglobin (NiHb) and myoglobin (NiMb) and model Ni porphyrins have been investigated by Soret-resonance Raman difference spectroscopy. Two sets of frequencies for the oxidation-state and core-size marker lines in the region from 1300 to 1700 cm-1 indicate two distinct sites in NiHb. Only one of these sites is evident in the Raman spectra of NiMb. This result is consistent with the UV-visible absorption spectrum of NiHb, which shows two Soret bands at 397 and 420 nm and one Soret at 424 nm for NiMb. Excitation at the blue Soret component of NiHb with 406.7-nm laser radiation preferentially enhances the set of Raman marker lines typical of Ni-protoporphyrin IX [Ni(ProtoP )] in noncoordinating solvents. The wavelength of the blue Soret component and the Raman spectrum indicate four-coordination for this site in NiHb. Laser excitation in the red Soret band enhances a set of lines whose frequencies are compatible with neither four- nor six-coordinate frequencies but are intermediate between the two. The red Soret band of the proteins is also considerably less red shifted than six-coordinate Ni-porphyrin models. These results suggest that Ni in the second site possesses a single axial ligand. Raman spectra of 64Ni-reconstituted and natural abundance Ni-reconstituted hemoglobins, obtained simultaneously in a Raman difference spectrometer, have identified the Ni-ligand stretch at 236 cm-1. The line shifts to 229 cm-1 for the 64Ni-reconstituted Hb. For a pure Ni-ligand stretch a 10-cm-1 shift would be predicted.(ABSTRACT TRUNCATED AT 250 WORDS)

Resonance Raman spectroscopic studies in copper reconstituted and hybrid hemoglobins: Probe into subunit heterogeneity

“…This band ( 4 ) is well known as the -charge density marker band of iron porphyrins, heme proteins, metalloporphyrins, and metal reconstituted hemoglobins. 16,20,22,24,29 A comparison of the high frequency Raman spectrum of CuHb at both 413.1-and 406.7-nm excitation wavelengths indicate that the core and oxidation state marker bands are consistent with the earlier reported 4-and 5-coordinate porphyrin systems. 16,24,29 Note here that the 4-and 5-coordinate Raman marker bands in CuHb are not clearly separated out as in the case of NiHb.…”

“…16,20,22,24,29 A comparison of the high frequency Raman spectrum of CuHb at both 413.1-and 406.7-nm excitation wavelengths indicate that the core and oxidation state marker bands are consistent with the earlier reported 4-and 5-coordinate porphyrin systems. 16,24,29 Note here that the 4-and 5-coordinate Raman marker bands in CuHb are not clearly separated out as in the case of NiHb. 16 This becomes obvious because even in the 4-and 5-coordinate model Cu porphyrins 24 these bands are indistinguishable with respect to their position.…”

“…Figure 1(a,b) shows the high frequency RR spectra of CuHb at 413.1-and 406.7-nm excitation wavelengths, respectively. The oxidation and core size marker bands ( 4 , 3 , 2 , and 10 ) are consistent with the model 4-and 5-coordinate copper porphyrins 16,24,29 (Table I). An interesting feature is the appearance of an additional 10 band at 1640 cm Ϫ1 that is exclusively present at 406.7-nm excitation and absent at 413.1-nm excitation.…”

“…16,24,29 Note here that the 4-and 5-coordinate Raman marker bands in CuHb are not clearly separated out as in the case of NiHb. 16 This becomes obvious because even in the 4-and 5-coordinate model Cu porphyrins 24 these bands are indistinguishable with respect to their position. The reason for this could be a smaller difference in the Soret band maxima of 4-and 5-coordinate forms of Cu systems, unlike in Ni reconstituted model systems.…”

“…In addition, UV-visible spectra of Cu and Ni Hbs show a split Soret band, 11,15 which is consistent with the presence of two metal ion environments. Shelnutt and coworkers subsequently demonstrated that nickel reconstituted Hb (NiHb) exhibits both 4 and 5 coordination 16 as revealed by resonance Raman (RR) enhancement of marker lines at different excitations. Additional support of heterogeneity in the metal ion coordination came from Shibayama et al by means of NMR studies 17 where it was shown that Ni(II) also exhibits a paramagnetic S ϭ 1 state (in addition to S ϭ 0) due to the weak axial ligation from the proximal histidine satisfying the 5th coordination.…”