Molecular Stability of he Philly (α₂ β₂^{35 (C1) Tyr → Phe})

Abstract

The relationship of hemoglobin stability to ligand state as defined by heat and mechanical shaking tests

The molecular stability of Hb Philly (α₂ β₂^{35 (C1) Tyr→Phe}) with different ligand states was compared with that of Hb A and Hb S using mechanical shaking and heat stability tests. The rates of mechanical denaturation of the oxy-forms of these hemoglobins decreased in the order of Hb S, Hb Philly, and Hb A, with relative ratios of 9.5: 5.6: 1.0. Upon oxidation to the met-forms, Hb Philly became mechanically most unstable, with ratios of 13.3: 23.0: 1.8, respectively. The deoxy-forms of Hb A and Hb S were very stable, while that of Hb Philly was as unstable as the oxy-form. The addition of inositol hexaphosphate (IHP) to deoxy-Hb Philly stabilized the molecules. Since IHP restores the cooperative oxygen binding of Hb Philly, deoxy-Hb Philly appears to combine with IHP to change the quaternary structure required for cooperative oxygen binding and for stabilization of the molecule.

Heat stability tests on Hb Philly showed that the oxy- and met-forms were slightly more unstable than those of Hb A, while the deoxy- and carbonmonoxy-forms were as stable as those of Hb A. Results for heat stability tests showed that hemoglobin molecules are stabilized when converted to the deoxy- or carbonmonoxy-forms so that oxidation of hemoglobin into the met-form is prevented. These results indicate that the stability of hemoglobin depends highly on its ligand state, and that the stability of various ligand forms should be tested when hemoglobin stability is investigated.