Quaternary structure and spin-state transition in azide methemoglobin A

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Abstract
The temperature-dependent UV and visible absorption changes of human azide methemoglobin with and without inositol hexaphosphate (IHP) were examined in a 4.degree.-35.degree. C range. The 537-nm absorption change of IHP-free Hb was about 1.2-fold larger than that of IHP-bound Hb. The data were analyzed by considering the thermal spin equilibrium within the R and T conformers and the quaternary equilibrium between the 2 conformers. The spin equilibrium analysis suggested that the T conformer has a larger high-spin content than the R conformer. The quaternary equilibrium analysis showed that the T conformer is more populated at lower temperature. The thermodynamic values for the quaternary equilibrium were determined to be .DELTA.H = -13.3 kcal/mol and .DELTA.S = -47.6 eu [entropy unit]. The large negative .DELTA.H and .DELTA.S values were compensated for each other to give a small energy difference between the two quaternary states, e.g., .DELTA.G4 = 670 cal/mol of tetramer at 20.degree. C. The coincidence of the temperature-dependent IHP-induced changes in the visible and UV absorptions of heme and aromatic chromophores at the subunit boundaries suggested that the quaternary transition energy is not localized at heme moiety. The reverse temperature dependence of the T conformer fraction as compared with the high-spin fraction of heme Fe was interpreted as indicating that the appearance of the T state is not directly coupled with an increase in the strain of Fe-N(F8 His) linkage in azide methemoglobin A.