We have studied the effect of the P6-inositol (IHP)-induced change from the quaternary oxy (R) to the deoxy (T) structure in derivatives of human, trout IV, and carp methemoglobins. Addition of IHP to human fluoroand aquomethemoglobin leads to the appearance of the slowly exchanging proton resonance at about -10 ppm from HDO diagnostic of the T structure. This experiment, and the crystallization of aquomethemoglobin + IHP by G. Fermi & M. F. Perutz ((1977) J. Mol. Biol. 114, 421) confirmed that the spectral change in the UV which IHP induces in these compounds can be used as a reliable indicator of the R-*-T transition. Judged by this spectral change, IHP converts all derivatives of carp hemoglobin from the R to the T structure. The pH at which the midpoint of the IHP-induced transition occurs increases with rising spin, being lowest in cyano, intermediate in azido, and highest in thiocyanate and aquomethemoglobin of carp. Conversely the replacement of water by fluoride or thiocyanate as the sixth ligand is unaffected by IHP because all three derivatives are predominantly high spin, but the affinity of azide for carp aquomethemoglobin is reduced 2.7-fold and that of cyanide 3.3-fold by IHP, corresponding to changes in the free energy of binding of 600 and 700 cal/mol heme. Conversion to the T structure of all carp methemoglobin derivatives except the cyanide one is accompanied by large changes in the visible absorption spectra, the most spectacular being that of the nitrite derivative whose color is changed from red to brown. IHP converts all human methemoglobin derivtleme-heme interaction arises from an equilibrium between states which differ in the tertiary structure of the a and 0 subunits and in their quaternary structure in the tetramer. This equilibrium is linked to the stereochemistry at the heme. In deoxyhemoglobin where the tense (T) quaternary structure is dominant, the heme irons are five coordinated and high spin.

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