Quaternary structures of intermediately ligated human hemoglobin a and influences from strong allosteric effectors: resonance Raman investigation.

The Fe-histidine stretching (nu(Fe-His)) frequency was determined for deoxy subunits of intermediately ligated human hemoglobin A in equilibrium and CO-photodissociated picosecond transient species in the presence and absence of strong allosteric effectors like inositol(hexakis)phosphate, bezafibrate, and 2,3-bisphosphoglycerate. The nu(Fe-His) frequency of deoxyHb A was unaltered by the effectors. The T-to-R transition occurred around m = 2-3 in the absence of effectors but m > 3.5 in their presence, where m is the average number of ligands bound to Hb and was determined from the intensity of the nu(4) band measured in the same experiment. The alpha1-beta2 subunit contacts revealed by ultraviolet resonance Raman spectra, which were distinctly different between the T and R states, remained unchanged by the effectors. This observation would solve the recent discrepancy that the strong effectors remove the cooperativity of oxygen binding in the low-affinity limit, whereas the (1)H NMR spectrum of fully ligated form exhibits the pattern of the R state.