MCD and MLD Studies of Cytochrome Oxidase and other Hemoproteins

细胞色素氧化酶和其他血红素蛋白的 MCD 和 MLD 研究

• 批准号: 9506817
• 负责人: James Peterson
• 金额: $ 5万
• 依托单位: University of Alabama Tuscaloosa
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-08-01 至 1996-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9506817&HistoricalAwards=false
• 关键词: MCD; MLD; Studies; Cytochrome; Oxidase

9506817 Peterson This research will develop magnetic linear dichroism (MLD) as a valuable tool for the study of cytochrome c oxidase and some other metalloproteins. MLD is not sensitive to heme a or CuA(which give rise to features that dominate the magnetic circular dichroism MCD spectra of most cytochrome c oxidase derivatives one would like to study) but is sensitive to some states in which the heme a3-CUB binuclear pair can be prepared. Consequently, it is expected that the site of interest can be observed free of spectral interference from the other chromophores using the MLD approach. The major limitation associated with magneto-optical measurements on hemoproteins in general has been that the MCD signals derived from species having a low spin ferric (S = 1/2) ground-state are the most intense and can therefore dominate much of the spectrum. Such hemes tend to be simply electron transfer centers and not directly involved in substrate binding. Since for most purposes we are primarily concerned with the latter, this represents something of a problem. Nearly always, it is possible to prepare one or more interesting derivatives where the electron transfer centers are low-spin ferrous (S = O) and the substrate binding heme has some paramagnetic ground-state (S =2 usually). In these circumstances, only the active site heme has a readily measurable MCD signal. Unfortunately, there invariably remain a number of derivatives which could usefully be studied by MCD spectroscopy if it were not for small amounts of residual low-spin ferric hemes dominating the spectrum. This tends to be the rule rather than the exception and is definitely the most serious source of spectral interference associated with the technique. MLD spectroscopy represents something of a breakthrough in this regard, since it is not sensitive to either low-sin ferric, or ferrous, systems; but is likely to prove particularly useful for studying integer spin paramagnets. The MCD instrumentatio n now on-line at The University of Alabama operates between 1.2 and 300 K, O to 7 tesla, over the wavelength range 350 to 3000 nm. No other single instrument has these capabilities. Also, unlike other MCD instruments, it has been constructed entirely by American companies. The planned addition of the MLD option will represent a further unique feature in a single instrument. This equipment arguably represents something of a national resource. %%% This research will develop the technique known as magnetic linear dichroism (MLD) as a tool for the study of hemoprotein and other samples which have proved difficult to investigate by established methods. Presently, there seem to be no useful MLD data reported for any metalloprotein samples.The information obtained will assist in the elucidation of the mechanism by which certain metalloenzymes function and is, therefore, of some fundamental significance to biochemistry. The planned addition of the MLD option to the magnetic circular dichroism (MCD) instrumentation now on-line at The University of Alabama will represent a further unique feature in a single instrument and, arguably, represents something of a national resource. ***

9506817 Peterson本研究将发展磁性线性二色性（MLD）作为研究细胞色素c氧化酶和其他金属蛋白的有价值的工具。MLD对血红素a或CuA不敏感（这引起主导磁性圆二色性的特征 MCD光谱的大多数细胞色素c氧化酶衍生物的人想研究），但敏感的一些国家，其中血红素a3-CUB双核对可以准备。因此，预期使用MLD方法可以观察到感兴趣的位点不受其他发色团的光谱干扰。 一般而言，与血红素蛋白的磁光测量相关的主要限制是来自具有低自旋铁（S = 1/2）基态的物种的MCD信号是最强的，因此可以支配大部分光谱。 这种血红素往往只是电子转移中心，不直接参与底物结合。由于在大多数情况下，我们主要关心的是后者，这就代表了一个问题。几乎总是可以制备一个或多个有趣的衍生物，其中电子转移中心是低自旋亚铁（S = O），并且底物结合血红素具有一些顺磁性基态（通常S =2）。在这些情况下，只有活性位点血红素具有容易测量的MCD信号。 不幸的是，总是有一些衍生物，可以有效地研究MCD光谱，如果它不是少量的残留低自旋铁血红素占主导地位的频谱。这往往是规则而不是例外，并且肯定是与该技术相关的光谱干扰的最严重来源。 MLD光谱在这方面代表了某种突破，因为它对低sin铁或亚铁系统不敏感;但可能被证明对研究整数自旋顺磁体特别有用。 目前在亚拉巴马大学联机的MCD仪器在1.2至300 K、0至7特斯拉、350至3000 nm波长范围内工作。没有其他单一的工具具有这些能力。此外，与其他MCD仪器不同，它完全由美国公司建造。计划增加的MLD选项将是单一仪器的另一个独特功能。这些设备可以说是某种国家资源。 本研究将开发被称为磁性线性二色性（MLD）的技术，作为研究血红素蛋白和其他已被证明难以用现有方法研究的样品的工具。目前，似乎还没有任何有用的MLD数据的金属蛋白样品的报告，所获得的信息将有助于阐明某些金属酶的功能机制，因此，是一些基本的生物化学意义。 计划增加的MLD选项的磁性圆二色性（MCD）仪器，现在在线在亚拉巴马大学将代表一个单一的仪器进一步独特的功能，可以说，代表了一些国家的资源。 ***