Resonance Raman studies of heme structural differences in subunits of deoxy hemoglobin

Low frequency resonance Raman (RR) spectra are reported for deoxy hemoglobi n (Hb), its isolated subunits, its analogue bearing methine-deuterated heme s in all four subunits (Hb-d(4)), and the hybrids bearing the deuterated he me in only one type of subunit, which are [alpha(d4)beta(h4)](2) and [alpha (h4)beta(d4)](2). Analyzed collectively, the spectra reveal subunit-specifi c modes that conclusively document subtle differences in structure for the heme prosthetic groups in the two types of subunits within the intact tetra mer. Not surprisingly, the most significant spectral differences are observ ed in the beta(7) mode that has a major contribution from out of plane bend ing of the methine carbons, a distortion that is believed to relieve strain in the high-spin heme prosthetic groups. The results provide convincing ev idence for the utility of selectively labeled hemoglobin hybrids in unravel ing the separate subunit contributions to the RR spectra of Hb and its vari ous derivatives and for thereby detecting slight structural differences in the subunits. (C) 2000 John Wiley & Sons, Inc.