Significance of beta116 His (G18) at alpha1beta1 contact sites for alphabeta assembly and autoxidation of hemoglobin.
Significance of beta116 His (G18) at alpha1beta1 contact sites for alphabeta assembly and autoxidation of hemoglobin.
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α1β1 接触位点的 beta116 His (G18) 对 Alphata 组装和血红蛋白自动氧化的意义。
DOI：
10.1021/bi030095s
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发表时间：
2003
期刊：
影响因子：
--
通讯作者：
Surrey,Saul
中科院分区：
文献类型：
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作者：
Adachi,Kazuhiko;Yang,Yi;Lakka,Vinaysagar;Wehrli,Suzanne;Reddy,KondaS;Surrey,Saul
杂四聚体相互作用位点在血红蛋白组装和自氧化中的作用尚不清楚。用重组β116Hisγ116HisγIle和→112Cys→Thr，116His→Ile和β112Cys→Thr，116His→Ilechains研究了Hb116His(G-18)和Hb116Ile分别位于α和β1β1或α1γ1相互作用位点之一对β和γ链形成同源二聚体和与α链在体外组装形成人Hb A和Hb F的重要性。尽管β链(例如116His)处于单体/四聚体平衡状态，但β116Aspchains仅显示单体形成。相反，β116Ile和β112Thr，116Ilechains显示出同源二聚体和同源四聚体形成，类似于含有116Ile的γ-珠蛋白链。β116Ileorβ112Thr、116Ilechain与α链的体外组装速度比正常β-珠蛋白链慢340倍，而β116Aspchains促进组装。这些结果还表明，非α链上G-18位的氨基酸疏水性在同源四聚体、二聚体和单体的形成中起着关键作用，而单体又在与α链组装形成Hb A和Hb F的过程中起着关键作用。这些结果还表明，γ-珠蛋白链的稳定二聚体不能在体内发生，因为这会抑制与α链的结合形成Hb F。还通过使用含有这些变异的Hb四聚体的重组Hb四聚体评估β116His(G-18)在异四聚体诱导的与血红蛋白中氧的键的稳定中的作用。α2β2116Asps和α2β2116Iletetramers的自氧化速率呈两相动力学，其中α链氧化速率较快，β链异构体的自氧化速率较慢，其速率是正常β-珠蛋白链的1.5倍。此外，这两个血红蛋白变体四聚体的血红素区域的核磁共振谱显示了与Hb A不同的共振峰，但α2β2116His→Aspandα2β2116His→Ile的氧结合特性与Hb A相比略有变化。这些结果表明，β116氨基酸(G18)不仅在稳定α1β1相互作用方面起关键作用，而且在抑制Hb氧化方面起着关键作用。然而，氧和血红素之间键的稳定可能不依赖于α1β1相互作用的稳定。β链与α链组装后，三级结构的变化可能会导致β链上的血红素区发生变化，从而影响DNA链上氧结合的稳定性。
The role of heterotetramer interaction sites in assembly and autoxidation of hemoglobin is not clear. The importance of β116His(G-18) and γ116Ileat one of the α1β1 or α1γ1 interaction sites for homo-dimer formation and assembly in vitro of β and γ chains, respectively, with α chains to form human Hb A and Hb F was assessed using recombinant β116His→Asp, β116His→Ile, and β112Cys→Thr,116His→Ilechains. Even though β chains (e.g., 116 His) are in monomer/tetramer equilibrium, β116Aspchains showed only monomer formation. In contrast, β116Ileand β112Thr,116Ilechains showed homodimer and homotetramer formation like γ-globin chains which contain 116 Ile. Assembly rates in vitro of β116Ileor β112Thr,116Ilechains with α chains were 340-fold slower, while β116Aspchains promoted assembly compared to normal β-globin chains. These results indicate that amino acid hydrophobicity at the G-18 position in non-α chains plays a key role in homotetramer, dimer, and monomer formation, which in turn plays a critical role in assembly with α chains to form Hb A and Hb F. These results also suggest that stable dimer formation of γ-globin chains must not occur in vivo,since this would inhibit association with α chains to form Hb F. The role of β116His(G-18) in heterotetramer-induced stabilization of the bond with oxygen in hemoglobin was also assessed by evaluating autoxidation rates using recombinant Hb tetramers containing these variant globin chains. Autoxidation rates of α2β2116Aspand α2β2116Iletetramers showed biphasic kinetics with the faster rate due to α chain oxidation and the slower to the β chain variants whose rates were 1.5-fold faster than that of normal β-globin chains. In addition, NMR spectra of the heme area of these two hemoglobin variant tetramers showed similar resonance peaks, which are different from those of Hb A. Oxygen-binding properties of α2β2116His→Aspand α2β2116His→Ile, however, showed slight alteration compared to Hb A. These results suggest that the β116 amino acid (G18) plays a critical role in not only stabilizing α1β1 interactions but also in inhibiting hemoglobin oxidation. However, stabilization of the bonds between oxygen and heme may not be dependent on stabilization of α1β1 interactions. Tertiary structural changes may lead to changes in the heme region in β chains after assembly with α chains, which could influence stability of dioxygen binding of β chains.