Structure and ligand binding properties of myoglobins reconstituted with monodepropionated heme: Functional role of each heme propionate side Chain
Structure and ligand binding properties of myoglobins reconstituted with monodepropionated heme: Functional role of each heme propionate side Chain
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DOI：
10.1021/bi7007068
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发表时间：
2007-08-21
期刊：
影响因子：
2.9
通讯作者：
Hayashi, Takashi
中科院分区：
文献类型：
--
作者：
Harada, Katsuyoshi;Makino, Masatomo;Hayashi, Takashi
在抹香鲸肌红蛋白中，连接在血红素骨架6和7位的两条血红素丙酸侧链分别与Arg45和Ser92相连。为了评估每个丙酸的作用，制备了两种单腿血红素，6-脱丙基和7-去丙基原血红素，并将其插入去肌红蛋白中，得到两种重组蛋白。重组肌红蛋白的结构数据通过1.1-1.4A分辨率的X射线结晶学分析和共振拉曼光谱获得。结果发现，6-丙酸的缺乏减少了末端位置的氢键数量，并明显改变了Arg45残基的位置，从而破坏了Arg45-Asp60的相互作用。相反，去除7-丙酸并不会导致远端和近端残基的显著结构变化。然而，共振拉曼研究表明，与天然血红素相比，蛋白质与7-去丙基原血红素的His93-Fe键的配位键强度略有增加。重组蛋白与单腿血红素的O-2和CO配体结合研究为了解每个丙酸的功能作用提供了重要的见解。与其他重组和天然蛋白质相比，缺乏6-丙酸可加速O-2解离约3倍。与蛋白质与天然血红素的亲和力相比，缺少7-丙酸使CO亲和力提高了2倍。这些结果表明，6-丙酸对结合的O-2有明显的稳定作用，而7-丙酸在调节Fe-His键方面起着重要作用。
Two heme propionate side chains, which are attached at the 6 and 7 positions of the heme framework, are linked with Arg45 and Ser92, respectively, in sperm whale myoglobin. To evaluate the role of each propionate, two kinds of one-legged hemins, 6-depropionated and 7-depropionated protohemins, were prepared and inserted into the apomyoglobin to yield two reconstituted proteins. Structural data of the reconstituted myoglobins were obtained via an X-ray crystallographic analysis at a resolution of 1.1-1.4 A and resonance Raman spectroscopy. It was found that the lack of the 6-propionate reduces the number of hydrogen bonds in the distal site and clearly changes the position of the Arg45 residue with the disrupting Arg45-Asp60 interaction. In contrast, the removal of the 7-propionate does not cause a significant structural change in the residues of the distal and proximal sites. However, the resonance Raman studies suggested that the coordination bond strength of the His93-Fe bond for the protein with the 7-depropionated protoheme slightly increases compared to that for the protein with the native heme. The O-2 and CO ligand binding studies for the reconstituted proteins with the one-legged hemes provide an important insight into the functional role of each propionate. The lack of the 6-propionate accelerates the O-2 dissociation by ca. 3-fold compared to those of the other reconstituted and native proteins. The lack of the 7-propionate enhances the CO affinity by 2-fold compared to that of the protein with the native heme. These results indicate that the 6-propionate clearly contributes to the stabilization of the bound O-2, whereas the 7-propionate plays an important role in the regulation of the Fe-His bond.