Resonance Raman and Electron Paramagnetic Resonance Studies of the Quaternary Structure Change in Carp Hemoglobin. Sensitivity of These Spectroscopic Probes to Heme Strain

It is possible to induce a change between T and R quaternary conformations of carp (Cyprinus carpio) hemoglobin merely by changing the pH and inositol hexaphosphate concentration. This provides a unique opportunity to examine the properties of liganded and of unliganded hemes in both low-and high-affinity conformations by resonance Raman (RR) and electron paramagnetic resonance (EPR) spectroscopies. The change in quaternary structure causes no shifts in the heme RR frequencies of the unliganded Hb, or of either high-or low-spin liganded derivatives. However, the induced structure change does alter the high-spin/low-spin equilibrium of azidomet-Hb from 10/90% to 45/55% and changes do occur in the EPR parameters of the low-spin liganded derivatives. These results have led us to examine the relative sensitivities of the two spectroscopic probes to small deformations (strains) in the heme geometry. For a low-spin six-coordinate heme, RR is predicted to be insensitive to small out of plane motions of the metal ion, while EPR is quite sensitive to coordination-geometry deformations. RR might be sensitive to deformations of a high-spin heme; EPR is not.