The heme-regulatory motifs of heme oxygenase-2 contribute to the transfer of heme to the catalytic site for degradation
The heme-regulatory motifs of heme oxygenase-2 contribute to the transfer of heme to the catalytic site for degradation
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DOI：
10.1074/jbc.ra120.012803
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发表时间：
2020-04-17
影响因子：
4.8
通讯作者：
Ragsdale, Stephen W.
中科院分区：
文献类型：
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作者：
Fleischhacker, Angela S.;Gunawan, Amanda L.;Ragsdale, Stephen W.
血红素调控基序（HRMs）存在于许多参与多种生物功能的蛋白质中。人血红素加氧酶-2 （HO2）的c端尾部区域包含两个HRMs，其半胱氨酸残基形成二硫键；当还原后，这些半胱氨酸可以结合Fe3+-血红素。血红素与HRMs的结合独立于蛋白质核心的HO2催化活性位点发生，血红素以高亲和力结合并降解为胆绿素。在这里，我们描述了可逆的，蛋白质介导血红素在HRMs和HO2核心之间的转移。利用氢-氘交换(HDX)-质谱法监测HO2在有Fe3+-血红素与HRMs和核心结合时和没有Fe3+-血红素结合时的动力学，我们仅在催化循环的一种状态下检测到催化核心的构象变化。当Fe3+血红素与HRMs结合时，核心处于载脂蛋白状态。这些构象变化与血红素在结合位点之间的转移一致。事实上，我们观察到hrm结合的Fe3+血红素通过核心和跨越hrm尾部的构建体的独立表达或在核心的血红素单次翻转后转移到载脂蛋白核心。此外，我们观察到血红素从核心转移到HRMs，以及血红素在核心和HRMs之间的平衡。因此，我们提出了一个Fe3+-血红素转移模型，其中hrm结合的血红素很容易转移到催化位点进行降解以促进周转，但也可以在位点之间保持平衡以维持血红素稳态。
Heme-regulatory motifs (HRMs) are present in many proteins that are involved in diverse biological functions. The C-terminal tail region of human heme oxygenase-2 (HO2) contains two HRMs whose cysteine residues form a disulfide bond; when reduced, these cysteines are available to bind Fe3+-heme. Heme binding to the HRMs occurs independently of the HO2 catalytic active site in the core of the protein, where heme binds with high affinity and is degraded to biliverdin. Here, we describe the reversible, protein-mediated transfer of heme between the HRMs and the HO2 core. Using hydrogen-deuterium exchange (HDX)-MS to monitor the dynamics of HO2 with and without Fe3+-heme bound to the HRMs and to the core, we detected conformational changes in the catalytic core only in one state of the catalytic cycle?when Fe3+-heme is bound to the HRMs and the core is in the apo state. These conformational changes were consistent with transfer of heme between binding sites. Indeed, we observed that HRM-bound Fe3+-heme is transferred to the apo-core either upon independent expression of the core and of a construct spanning the HRM-containing tail or after a single turnover of heme at the core. Moreover, we observed transfer of heme from the core to the HRMs and equilibration of heme between the core and HRMs. We therefore propose an Fe3+-heme transfer model in which HRM-bound heme is readily transferred to the catalytic site for degradation to facilitate turnover but can also equilibrate between the sites to maintain heme homeostasis.