Structural and functional studies of hemoglobin Poissy alpha 2 beta 2(56) (D7) Gly----Arg and 86 (F2) Ala----Pro.

Abstract

Hemoglobin Poissy alpha 2 beta 2(56) (D7) Gly----Arg and 86 (F2) Ala----Pro, is a new variant of the beta chain with two substitutions within the second exon of the corresponding gene. The electrophoretic mobilities are identical to those of Hb Hamadan alpha 2 beta 2(56) (D7) Gly----Arg as is the fingerprint of the tryptic hydrolysate of the two abnormal beta chains. The second substitution beta 86 Ala----Pro was detected by high-pressure liquid chromatography. Hb Poissy has a threefold increase in oxygen affinity with low Hill coefficient and diminished Bohr effect, which are restored to normal upon addition of 2,3-bisphosphoglycerate. Since the functional properties of Hb Hamadan (beta 56 Gly----Arg) have been described as normal, the abnormal function of Hb Poissy may be attributed to the beta 86 (F2) Ala----Pro substitution. Hb Poissy exhibits a mild instability and a greater reactivity of the thiol groups of the beta 93 (F9) Cys residues in the deoxy form than does Hb A. The oxidation rate of Hb Poissy is biphasic indicating a large inequivalence between the alpha and beta hemes. Thereafter NMR studies demonstrated that the beta 86 Ala----Pro substitution produces a displacement of the F helix closer to the heme plane and a large increase in the dynamic fluctuations of the tertiary structure on the proximal side of the beta hemes. These results lead to the conclusion that the beta 86 Ala----Pro substitution produces a destabilization of the F helix extending downwards to the FG corner and altering both the beta hemes and the alpha 1 beta 2 contacts.