Spectroscopic studies of the nature of ligand bonding in carbonmonoxyhemoglobins: evidence of a specific function for histidine-E7 from infrared and nuclear magnetic resonance intensities.

Infrared spectra of carbon monoxide ligated hemoglobins from human, horse, and rabbit donors have been examined. A single vibrational frequency at 1951 cm-1 is observed for CO bound to the heme in horse and human hemoglobins. Studies of the isolated alpha-CO and beta-CO subunits of human hemoglobin reveal that the observation of a single frequency in the intact tetramer is the result of a superposition of the alpha-CO and beta-CO vibrational frequencies. The apparent integrated absorption intensities of these CO vibrations are shown both to have values of 1.0 X 10(5)M-1cm-2 within experimental error. For rabbit CO-Hb two vibrational frequencies appear (Caughey, W. S., et al. (1973) Fed. Proc., Fed. Am. Soc. Exp. Biol. 32, 552) and are assigned to CO bound to the beta (1951 cm-1) and alpha(1928 cm-1) subunits within the intact tetramer. The beta-CO subunit exhibits both frequency and intensity similarities with horse and human hemoglobins. The rabbit alpha-CO subunit, however, exhibits a markedly lower frequency and much smaller intensity compared with the other CO-hemoglobins. These data are interpreted in terms of a specific role for the distal histidine (E7) in rabbit alpha subunits, in which this histidine functions as a nucleophilic donor to coordinated CO.