Effects of organic phosphates on oxygen equilibria and kinetics of SH reaction in feline hemoglobins

Abstract

A recent report indicates that the response of frog and tadpole hemoglobin to inositol hexaphosphate (IHP) varies from observations made on hemoglobins of other species ( Araki, T., Kajita, A., and Shukuya, R. (1971) Biochem Biophys. Res. Commun. 43, 1179). Since frog hemoglobins are known to be acetylated, investigation of the effects of IHP on the oxygen affinity of the acetylated (HbB) and nonacetylated (HbA) feline hemoglobins was of interest. The effect of IHP was of particular concern since it was previously shown that the oxygen affinity and SH reactivity of HbA is lowered by 2,3-diphosphoglycerate (2,3-DPG) and ATP while that of HbB is unaffected ( Taketa, F., Mauk, A. G., and Lessard, J. L. (1971) J. Biol. Chem. 246: 4471). In contrast to the relatively weak effect of 2,3-DPG on the oxygen affinity and SH reactivity of cat HbA compared with its effect on human HbA, the effect of IHP on cat HbA is much greater and is comparable to its effect on human HbA. However, both phosphate compounds are ineffective with cat HbB; insensitivity of cat HbB presumably arises from the occurrence of N-acetylated β-chain amino termini as well as substitutions at or near histidine 143-β of this protein. Thus, properties of the two cat hemoglobins are sharply differentiated in the presence of IHP. Consequently the response of cat hemolyzates that contain a 1:1 mixture of HbA and HbB is such that the shape of the oxygen saturation curve changes drastically in the presence of this effector. Good correlation was found with respect to the effects of 2,3-DPG and IHP on the oxygen affinities and SH reactivities in the cat hemoglobins. The results suggest that the organic phosphates induce conformational changes in these proteins that stabilize the deoxy form of the protein.