ABSENCE OF HEME-LOCALIZED STRAIN IN T-STATE HEMOGLOBIN - INSENSITIVITY OF HEME-IMIDAZOLE RESONANCE RAMAN FREQUENCIES TO QUATERNARY STRUCTURE

Substitution of pentadeuterated 2-methylimidazole in (2-methylimidazole)-Fe(II)-protoporphyrin IX, a model complex for deoxyHb, shifts three bands in the low-frequency resonance Raman spectrum 380→373 cm-1, 348→345 cm-1, and 220→218 cm-1. The first of these is assigned primarily to Fe-imidazole stretching, and the other two are assigned to porphyrin deformation modes with substantial Fe-pyrrole stretching contributions. The three bands are observed in deoxyHb and Mb. The Fe-pyrrole modes are at essentially the same frequencies in the two proteins, but the Fe-imidazole mode is 6 cm-1 lower in deoxyHb than Mb, implying a slight alteration in the heme-imidazole linkage. No change greater than 2 cm-1 is observed when Hb Kempsey is switched from the R to the T state. This observation places an upper limit on the energy stored in the Fe-imidazole bond of T state deoxyHb, which is estimated to be <0.2 kcal/mol (<836.8J/mol).