Polymerization of three hemoglobin A2 variants containing Val and inhibition of hemoglobin S polymerization by hemoglobin A2

Abstract

To understand determinants for hemoglobin (Hb) stability and Hb A2 inhibition of Hb S polymerization, three Val56 Hb A2 variants (Hb A2 δE6V, Hb A2 δE6V,δQ87T, and Hb A2 δE6V,δA22E,δQ87T) were expressed in yeast, and stability to mechanical agitation and polymerization properties were assessed. Oxy forms of Hb A2 δE6V and Hb A2 δE6V, δQ87T were 2- and 1.6-fold, respectively, less stable than oxy-Hb S, while the stability of Hb A2 δE6V,δA22E,δQ87T was similar to that of Hb S, suggesting that Alaδ22 and Ginδ87 contribute to the surface hydrophobicity of Hb A2. Deoxy Hb A2 δE6V polymerized without a delay time, like deoxy Hb F γE6V, while deoxy Hb A2 δE6V, δQ87T and deoxy Hb A2 δE6V,δA22E,δQ87T polymerized after a delay time, like deoxy Hb S, suggesting that β87 Thr is required for the formation of nuclei. Deoxy Hb F γE6V, γQ87T showed no delay time and required a 3.5-fold higher concentration than deoxy Hb S for polymerization, suggesting that Thr effects on Val(δ6) Hb A2 and Valγ6 Hb F variants are different. Mixtures of deoxy Hb S/Hb A2 δE6V,δVQ87T polymerized, like deoxy Hb S, while polymerization of Hb S/Hb A2 δE6V mixtures was inhibited, like Hb S/Hb F γE6V mixtures. These results suggest α2 β(S) δ6 Val, 87 Thr hybrids and Hb A2 δE6V,δQ87T participate in Hb S nucleation, while only 50% of α2β(S)δ Val hybrids and none of the Hb A2 δE6V participate. These findings are in contrast to those of mixtures of Hb S with Hb F γE6V or Hb F γE6V,Q87T, which both inhibit Hb S polymerization. Our results also suggest participation in nucleation of some aα2β(S) hybrids in A2S mixtures but not α2β(S)γ hybrids in FS mixtures.