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首页> 外文期刊>Journal of Molecular Biology >Interspecies hybrid HbS: complete neutralization of Val6(beta)-dependent polymerization of human beta-chain by pig alpha-chains.
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Interspecies hybrid HbS: complete neutralization of Val6(beta)-dependent polymerization of human beta-chain by pig alpha-chains.

机译：种间杂种HbS：猪α-链完全中和人β-链的Val6β依赖性聚合。

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Interspecies hybrid HbS (alpha(2)(P)beta(2)(S)), has been assembled in vitro from pig alpha-globin and human beta(S)-chain. The alpha(2)(P)beta(2)(S) retains normal tetrameric structure (alpha(2)beta(2)) of human Hb and an O(2) affinity comparable to that of HbS in 50 mM Hepes buffer; but, its O(2) affinity is slightly higher than that of HbS in the presence of allosteric effectors (chloride, DPG and phosphate). The (1)H-NMR spectroscopy detected distinct differences between the heme environments and alpha(1)beta(1) interfaces of pig Hb and HbS, while their alpha(1)beta(2) interfaces appear very similar. The interspecies hybrid alpha(2)(H)beta(2)(P) resembles pig Hb; the pig beta-chain dictated the conformation of the heme environment of the human alpha-subunit, and to the alpha(1)beta(1) interfaces of the hybrid. In the alpha(2)(P)beta(2)(S) hybrid, beta(S)-chain dictated the conformation of human heme environment to the pig alpha-chain in the hybrid; but the conformation of alpha(1)beta(1) interface of this hybrid is close to, but not identical to that of HbS. On the other hand, the alpha(1)beta(2) interface conformation is identical to that of HbS. More important, the alpha(2)(P)beta(2)(S) does not polymerize when deoxygenated; pig alpha-chain completely neutralizes the beta(S)-chain dependent polymerization. The polymerization inhibitory propensity of pig alpha-chain is higher when it is present in the cis alpha(P)beta(S) dimer relative to that in a trans alpha(P)beta(A) dimer. The semisynthetically generated chimeric pig-human and human-pig alpha-chains by exchanging the alpha(1-30) segments of human and pig alpha-chains have established that the sequence differences of pig alpha(31-141) segment can also completely neutralize the polymerization. Comparison of the electrostatic potential energy landscape of the alpha-chain surfaces of HbS and alpha(2)(P)beta(2)(S) suggests that the differences in electrostatic potential energy surfaces on the alpha-chain of alpha(2)(P)beta(2)(S) relative to that in HbS, particularly the ones involving CD region, E-helix and EF-corner of pig alpha-chain are responsible for the polymerization neutralization activity. The pig and human-pig chimeric alpha-chains can serve as blueprints for the design of a new generation of variants of alpha-chain(s) suitable for the gene therapy of sickle cell disease. Copyright 2000 Academic Press.
机译：种间杂种HbS（alpha（2）（P）beta（2）（S）），已从猪的α-珠蛋白和人β（S）-链体外组装而成。 alpha（2）（P）beta（2）（S）保留人类Hb的正常四聚体结构（alpha（2）beta（2））和与HbS在50 mM Hepes缓冲液中可比的O（2）亲和力;但是，在存在变构效应物（氯化物，DPG和磷酸盐）的情况下，其O（2）亲和力略高于HbS。 （1）H-NMR光谱检测到血红素环境与猪Hb和HbS的alpha（1）beta（1）界面之间的明显差异，而它们的alpha（1）beta（2）界面看起来非常相似。种间杂种alpha（2）（H）beta（2）（P）类似于猪Hb;猪的β链决定了人类α-亚基血红素环境的构象，并决定了杂种的α（1）β（1）界面。在alpha（2）（P）beta（2）（S）杂种中，beta（S）链决定了人类血红素环境与杂种中猪α链的构象;但是此杂合体的alpha（1）beta（1）界面的构型与HbS接近但不完全相同。另一方面，alpha（1）beta（2）界面构型与HbS相同。更重要的是，脱氧时，alpha（2）（P）beta（2）（S）不会聚合;猪的α-链完全中和了β（S）-链依赖性聚合。相对于反式α（P）beta（A）二聚体，猪α-链的聚合抑制倾向更高。通过交换人和猪α-链的alpha（1-30）片段，半合成生成的猪人-猪和人-猪alpha链已经确定，猪alpha（31-141）片段的序列差异也可以完全中和聚合。 HbS和alpha（2）（P）beta（2）（S）的α链表面的静电势能图的比较表明，α（2）（相对于HbS中的P）beta（2）（S），尤其是涉及猪α链的CD区，E螺旋和EF角的HbS负责聚合中和活性。猪和人猪嵌合α-链可以作为设计适于镰状细胞疾病基因治疗的新一代α-链变体的蓝图。版权所有2000学术出版社。

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