Heme-regulated eIF-2α kinase purifies as a hemoprotein
Heme-regulated eIF-2α kinase purifies as a hemoprotein
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DOI：
10.1046/j.1432-1327.1998.2580820.x
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发表时间：
1998-12-01
期刊：
影响因子：
--
通讯作者：
Chen, JJ
中科院分区：
文献类型：
--
作者：
Chefalo, PJ;Oh, JH;Chen, JJ
网织红细胞中血红素的可获得性对蛋白质合成的调节是公认的。然而，血红素调节翻译起始的机制尚不清楚。在这项研究中，我们直接研究了血红素对同种血红素调节的eIF-2α激酶(HRI)的调节，该蛋白在血红素缺乏时被激活。我们发现HRI纯化为一种血红素蛋白，其特征Soret带位于424 nm处。其活性可被亚微摩尔浓度的氯化高铁血红素抑制，表观K值为0.5MU。均一的HRI为同源二聚体，与纯化的失活K199R HRI体外孵育不能抑制其活性。我们的结果表明，在HRI同源二聚体中存在两种不同类型的血红素结合SIR。血红素与第一位点的结合是稳定的，而第二位位的结合是血红素快速下调HRI活性的原因。这些结果表明，HRI结合了血红素，并作为血红素可用性的感受器，协调红系细胞中珠蛋白和血红素的平衡合成。
The regulation of protein synthesis by the availability of heme in reticulocytes is well established. However, the mechanism by which heme regulates translational initiation is not clear. In this study, we have examined the heme regulation directly on the homogeneous heme-regulated eIF-2 alpha kinase (HRI), which is activated during heme deficiency. We found that HRI purified as a hemoprotein with the characteristic Soret band of hemoprotein at 424 nm. This HRI was an active autokinase and eIF-2 alpha kinase, and its kinase activities were inhibited by submicromolar concentrations of hemin with an apparent K, of 0.5 mu M Homogeneous HRI was a homodimer, and its activities could not be inhibited by incubation with purified inactive K199R HRI in vitro. Our results suggest that there are two distinct types of heme-binding sires in the HRI homodimer. The binding of heme to the first site is stable, while the binding of heme to the second site is responsible for the rapid downregulation of HRI activity by heme, These results indicate that HRI binds heme and sen es as a sensor of the availability of heme to coordinate the balanced synthesis of globins and heme in erythroid cells.