Sickle cell disease (SCD) is an inherited blood disorder caused by a β globin gene mutation of hemoglobin (HbS). Polymerization of deoxyHbS and subsequent aggregation (into long fibers) is the primary molecular event which leads to red blood cell (RBC) sickling and ultimately hemolytic anemia. We have recently suggested that HbS oxidative toxicity may also contribute to SCD pathophysiology due to its defective pseudoperoxidase activity. As a consequence, a persistent higher oxidization ferryl heme is formed which irreversibly oxidizes “hotspot” residues (particularly βCys93) causing protein unfolding and subsequent heme loss. In this report we confirmed first, the allosteric effect of a newly developed reagent (Di(5-(2,3-dihydro-1,4-benzodioxin-2-yl)- 4H-1,2,4-triazol- 3-yl)disulfide (TD-1) on oxygen affinity within SS RBCs. There was a considerable left shift in oxygen equilibrium curves (OECs) representing treated SS cells. Under hypoxic conditions, TD-1 treatment of HbS resulted in approximately 200 sec increase in the delay time of HbS polymerization over the untreated HbS control. The effect of TD-1 binding to HbS was tested next on oxidative reactions by incrementally treating HbS with increasing hydrogen peroxide (H2O2) concentrations. Under these experimental conditions, ferryl levels were consistently reduced by approximately 35% in the presence of TD-1. Mass spectrometric analysis confirmed that upon binding to Cys93, TD-1 effectively blocked irreversible oxidation of this residue. In conclusion, TD-1 appears to shield βCys93, the end point of radical formation in HbS, and when coupled with its modification of oxygen affinity it may provide new therapeutic modalities for the treatment of SCD.

中文翻译：

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用具有双重变构和抗氧化作用的抗镰刀蛋白靶向血红蛋白S中的βCys93

镰状细胞病（SCD）是由血红蛋白（HbS）的β珠蛋白基因突变引起的遗传性血液病。脱氧HbS的聚合和随后的聚集（长纤维聚集）是导致红细胞（RBC）镰刀化并最终导致溶血性贫血的主要分子事件。最近，我们建议HbS的氧化毒性也可能由于其伪过氧化物酶活性缺陷而导致SCD病理生理。结果，形成了持久的较高氧化态的亚铁血红素，其不可逆地氧化了“热点”残基（尤其是βCys93），导致蛋白质解折叠并随后导致血红素丢失。在本报告中，我们首先确认了新开发的试剂（Di（5-（2,3-dihydro-1,4-benzodioxin-2-yl）-4H-1,2,4-triazol- 3-芳基二硫化物（TD-1）对SS RBC中的氧亲和力的影响。代表处理过的SS细胞的氧平衡曲线（OEC）有相当大的左移。在低氧条件下，HbS的TD-1处理导致HbS聚合的延迟时间比未处理的HbS对照大约增加200秒。接下来，通过用增加的过氧化氢（H2O2）浓度逐步处理HbS，来测试TD-1与HbS的结合对氧化反应的影响。在这些实验条件下，在存在TD-1的情况下，三聚氰胺水平持续降低约35％。质谱分析证实，与Cys93结合后，TD-1有效地阻止了该残基的不可逆氧化。总之，TD-1似乎能够屏蔽HbS中自由基形成的终点βCys93，