Polymerization of three hemoglobin A(2) variants containing Val(delta 6) and inhibition of hemoglobin S polymerization by hemoglobin A(2)

To understand determinants for hemoglobin (Hb) stability and Hb A(2) inhibition of Hb S polymerization, three Val(delta 6) Hb A(2) variants (Hb A(2) delta E6V, Hb A(2) delta E6V, delta Q87T, and Hb A(2) delta E6V, delta A22E, delta Q87T) were expressed in yeast, and stability to mechanical agitation and polymerization properties were assessed. Oxy forms of Hb A(2) delta E6V and Hb A(2) delta E6V, delta Q87T were 2- and 1.6-fold, respectively, less stable than oxy-Hb S, while the stability of Hb A(2) delta E6V, delta A22E, delta Q87T was similar to that of Hb S, suggesting that Ala(delta 22) and Gln(delta 87) contribute to the surface hydrophobicity of Rb A(2). Deoxy Hb A(2) delta E6V polymerized without a delay time, like deoxy Hb F gamma EGV, while deoxy Hb A(2) delta E6V, delta Q87T and deoxy Hb A(2) delta E6V, delta A22E, delta Q87T polymerized after a delay time, like deoxy Hb S, suggesting that beta 87 Thr is required for the formation of nuclei. Deoxy Hb F gamma E6V, gamma Q87T showed no delay time and required a 3.5-fold higher concentration than deoxy Hb S for polymerization, suggesting that Thr effects on Val(delta 6) Hb A(2) and Val(gamma 6) Hb F variants are different. Mixtures of deoxy Hb S/Hb A(2) delta E6V, delta Q87T polymerized, like deoxy Hb S, while polymerization of Hb S/Hb A(2) delta E6V mixtures was inhibited, like Hb S/Hb F gamma E6V mixtures. These results suggest alpha(2) beta(S) delta(6 Val, 87 Thr) hybrids and Hb A(2) delta E6V, delta 87T participate in Hb S nucleation, while only 50% of alpha(2) beta(S) delta(6Val) hybrids and none of the Hb A(2) delta E6V participate. These findings are in contrast to those of mixtures of Hb S with Hb F gamma E6V or Hb F gamma E6V, Q87T, which both inhibit Hb S polymerization. Our results also suggest participation in nucleation of some alpha(2) beta(S) delta hybrids in A(2)S mixtures but not alpha(2) beta(S) gamma hybrids in FS mixtures.