Interactions between heme d and heme b595 in quinol oxidase bd from Escherichia coli:: A photoselection study using femtosecond spectroscopy
Interactions between heme d and heme b595 in quinol oxidase bd from Escherichia coli:: A photoselection study using femtosecond spectroscopy
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DOI：
10.1021/bi0158019
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发表时间：
2002-02-05
期刊：
影响因子：
2.9
通讯作者：
Vos, MH
中科院分区：
文献类型：
--
作者：
Borisov, VB;Liebl, U;Vos, MH
飞秒光谱进行CO-配体（完全还原和混合价态）和O-2-配体的醌醇氧化酶bd从大肠杆菌。在CO光解离光谱中观察到血红素蛋白质的光学研究中前所未有的大量偏振效应，这意味着血红素d（二氢卟酚配体结合位点）和靠近的血红素B（595）之间在皮秒时间尺度上的相互作用;该一般结果与先前的工作完全一致[Vos，M. H、Borisov，V. B.，利布尔大学，Martin，J.- L.的，和Konstantinov，A. A.等人（2000）Proc. Acad. Sci. U.S.A.97，15541559]。在各向同性和各向异性偏振条件下获得的数据和额外的闪光光解纳秒实验的突变体细胞色素bd主要缺乏血红素b595允许属性的功能，在众所周知的，但不寻常的CO解离光谱的细胞色素bd个人血红素d和血红素b595转换。这使得它可以比较的光谱CO解离还原和混合价细胞色素bd在静态条件下，并在皮秒的时间尺度上比以前更详细。血红素d的CO结合/解离被证明会干扰亚铁血红素b595，导致中心位于435 nm附近的吸收带的诱导/损失。此外，在50 ps的CO光解诱导的吸收变化揭示了亚铁血红素b595相对于静态光谱的红移。没有证据表明从血红素d解离后CO与血红素b595的瞬时结合在皮秒时间范围内。发现当血红素B（595）被氧化而不是还原时，在< 15 ps的时间尺度上，血红素d的CO光解离产率减少了3倍以上。与其他已知的血红素蛋白相比，分子氧不能从混合价的细胞色素bd中光解离，这表明酶中的二血红素活性位点具有独特的结构和电子构型。
Femtosecond spectroscopy was performed on CO-liganded (fully reduced and mixed-valence states) and O-2-liganded quinol oxidase bd from Escherichia coli. Substantial polarization effects, unprecedented for optical studies of heme proteins, were observed in the CO photodissociation spectra, implying interactions between heme d (the chlorin ligand binding site) and the close-lying heme b(595) on the picosecond time scale; this general result is fully consistent with previous work [Vos, M. H., Borisov, V. B., Liebl, U., Martin, J.-L., and Konstantinov, A. A. (2000) Proc. Natl. Acad. Sci. U.S.A. 97, 15541559]. Analysis of the data obtained under isotropic and anisotropic polarization conditions and additional flash photolysis nanosecond experiments on a mutant of cytochrome bd mostly lacking heme b595 allow to attribute the features in the well-known but unusual CO dissociation spectrum of cytochrome bd to individual heme d and heme b595 transitions. This renders it possible to compare the spectra of CO dissociation from reduced and mixed-valence cytochrome bd under static conditions and on a picosecond time scale in much more detail than previously possible. CO binding/dissociation from heme d is shown to perturb ferrous heme b595, causing induction/loss of an absorption band centered at similar to435 nm. In addition, the CO photodissociation-induced absorption changes at 50 ps reveal a bathochromic shift of ferrous heme b595 relative to the static spectrum. No evidence for transient binding of CO to heme b595 after dissociation from heme d is found in the picosecond time range. The yield of CO photodissociation from heme d on a time scale of < 15 ps is found to be diminished more than 3-fold when heme b(595) is oxidized rather than reduced. In contrast to other known heme proteins, molecular oxygen cannot be photodissociated from the mixed-valence cytochrome bd at all, indicating a unique structural and electronic configuration of the diheme active site in the enzyme.