Adachi K, Pang J, Reddy L R, Bradley L E, Chen Q, Trifillis P, Schwartz E, Surrey S
Children's Hospital of Philadelphia, Division of Hematology, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104, USA.
J Biol Chem. 1996 Oct 4;271(40):24557-63. doi: 10.1074/jbc.271.40.24557.
To understand determinants for hemoglobin (Hb) stability and Hb A2 inhibition of Hb S polymerization, three Valdelta6 Hb A2 variants (Hb A2 deltaE6V, Hb A2 deltaE6V,deltaQ87T, and Hb A2 deltaE6V, deltaA22E,deltaQ87T) were expressed in yeast, and stability to mechanical agitation and polymerization properties were assessed. Oxy forms of Hb A2 deltaE6V and Hb A2 deltaE6V,deltaQ87T were 2- and 1.6-fold, respectively, less stable than oxy-Hb S, while the stability of Hb A2 deltaE6V,deltaA22E,deltaQ87T was similar to that of Hb S, suggesting that Aladelta22 and Glndelta87 contribute to the surface hydrophobicity of Hb A2. Deoxy Hb A2 deltaE6V polymerized without a delay time, like deoxy Hb F gammaE6V, while deoxy Hb A2 deltaE6V,deltaQ87T and deoxy Hb A2 deltaE6V,deltaA22E,deltaQ87T polymerized after a delay time, like deoxy Hb S, suggesting that beta87 Thr is required for the formation of nuclei. Deoxy Hb F gammaE6V,gammaQ87T showed no delay time and required a 3.5-fold higher concentration than deoxy Hb S for polymerization, suggesting that Thr effects on Valdelta6 Hb A2 and Valgamma6 Hb F variants are different. Mixtures of deoxy Hb S/Hb A2 deltaE6V,deltaQ87T polymerized, like deoxy Hb S, while polymerization of Hb S/Hb A2 deltaE6V mixtures was inhibited, like Hb S/Hb F gammaE6V mixtures. These results suggest alpha2betaSdelta6 Val, 87 Thr hybrids and Hb A2 deltaE6V,deltaQ87T participate in Hb S nucleation, while only 50% of alpha2betaSdelta6 Val hybrids and none of the Hb A2 deltaE6V participate. These findings are in contrast to those of mixtures of Hb S with Hb F gammaE6V or Hb F gammaE6V,Q87T, which both inhibit Hb S polymerization. Our results also suggest participation in nucleation of some alpha2betaSdelta hybrids in A2S mixtures but not alpha2betaSgamma hybrids in FS mixtures.
为了解血红蛋白（Hb）稳定性的决定因素以及Hb A2对Hb S聚合的抑制作用，在酵母中表达了三种Valδ6 Hb A2变体（Hb A2 δE6V、Hb A2 δE6V,δQ87T和Hb A2 δE6V,δA22E,δQ87T），并评估了其对机械搅拌的稳定性和聚合特性。Hb A2 δE6V和Hb A2 δE6V,δQ87T的氧合形式分别比氧合Hb S不稳定2倍和1.6倍，而Hb A2 δE6V,δA22E,δQ87T的稳定性与Hb S相似，这表明Alaδ22和Glnδ87有助于Hb A2的表面疏水性。脱氧Hb A2 δE6V像脱氧Hb F γE6V一样无延迟时间地聚合，而脱氧Hb A2 δE6V,δQ87T和脱氧Hb A2 δE6V,δA22E,δQ87T像脱氧Hb S一样在延迟时间后聚合，这表明β87位的苏氨酸是形成核所必需的。脱氧Hb F γE6V,γQ87T无延迟时间，且聚合所需浓度比脱氧Hb S高3.5倍，这表明苏氨酸对Valδ6 Hb A2和Valγ6 Hb F变体的影响不同。脱氧Hb S/Hb A2 δE6V,δQ87T的混合物像脱氧Hb S一样聚合，而Hb S/Hb A2 δE6V混合物的聚合像Hb S/Hb F γE6V混合物一样受到抑制。这些结果表明α2βSδ6 Val、87 Thr杂合体和Hb A2 δE6V,δQ87T参与Hb S成核，而只有50%的α2βSδ6 Val杂合体参与，Hb A2 δE6V则无参与。这些发现与Hb S与Hb F γE6V或Hb F γE6V,Q87T混合物的结果相反，后两者均抑制Hb S聚合。我们的结果还表明，一些α2βSδ杂合体参与A2S混合物的成核，但FS混合物中的α2βSγ杂合体不参与。
