Spatial dynamics of electron transport

生命依赖于能量转换的过程，在这个过程中，从阳光中获得的能量，或者从催化化学反应（如食物分子的分解）中获得的能量，被转换成可以用来为细胞的生物化学提供动力的能量。一些关键的生物能转换过程发生在膜上，涉及电子从供体到受体的传递，由光能（光合作用）或化学能（呼吸作用）提供动力。这些过程需要一组膜组分，包括蛋白质复合物和可以在膜中传输电子的小分子。所涉及的成分现在已经被很好地理解了，但它们在完整膜中的组织和相互作用却知之甚少。在某些情况下，电子传递还有其他可能的途径。电子传递途径受生理因素的调控，对细胞的生理有很强的影响。很明显，电子传递的途径一定在某种程度上取决于膜中电子传递组分的组织。然而，目前尚不清楚电子传递发生在什么长度尺度上（即电子在膜上从初始供体到最终受体的距离），以及在什么长度尺度上可能组织复合物以控制电子传递途径。了解这些问题的答案可以为我们提供强大的新工具来控制生物能量转换，例如，允许工程或生物适合更有效的生物燃料生产。我们使用一种蓝藻（一种光合细菌）作为模式生物。在蓝藻中，光合作用和呼吸作用都发生在细胞内称为类囊体膜的复杂膜系统中。我们最初的方法来理解在蓝藻电子传输路线的控制已经使用的技术，我们产生突变体，其中特定的电子传输蛋白融合到荧光蛋白。然后，我们可以使用荧光显微镜来观察活细胞中完整膜中电子传递蛋白的分布。荧光显微镜的一个缺点是它的分辨率相对较低，即我们无法看到分子尺度上电子传递组分的组织。然而，这项技术使我们能够观察到，在我们所能观察到的尺度上，电子传递复合物的分布是非常不均匀的。在某些条件下，特定的电子传递组分集中在膜上不同的斑块上。我们可以证明，配合物在斑块中的分布，或以其他方式，是受生理控制的，我们可以将这与电子传递的生理控制联系起来。我们在这个项目中的目标是更好地了解膜斑块是如何形成的，它们包含什么，它们是如何组织的，以及它们对电子传递的影响。最初，我们将使用当前荧光标记技术的扩展，以获得活细胞中斑块组成的更完整的图像。这将与研究相结合，以测试在膜中复合物分布受到生理控制的方式的假设。然后，我们将使用生化技术分离膜斑块并确定其全部组成，并结合使用电子显微镜获得膜中蛋白质复合物组织的更高分辨率信息。在研究结束时，我们希望能更好地理解膜中的电子传递是如何被控制的。我们希望我们对蓝藻的研究能够作为其他生物膜尺度上生物电子传递研究的范例，并为一般生物膜的组织和功能的控制提供新的思路。

Life depends on processes of energy conversion, in which energy obtained from sunlight, or from catalysing chemical reactions such as the breakdown of food molecules, is converted into energy in forms that can be used to power the biochemistry of the cell. Some key biological energy conversion processes take place in membranes and involves the transport of electrons from donors to acceptors, powered by either light (photosynthesis) or chemical energy (respiration). These processes require a set of membrane components including protein complexes and smaller molecules that can transport electrons in the membrane. The components involved are now well understood, but their organisation and interactions in the intact membrane are much less understood. In some cases there are alternative possible routes for electron transport. Electron transport routes are regulated by physiological factors, and have strong effects on the physiology of the cell. It seems clear that the pathways of electron transport must depend somehow on the organisation of the electron transport components in the membrane. However, it remains unclear on what length scales electron transport takes place (i.e. how far in the membrane do electrons travel from their initial donor to their final acceptor) and on what length scales complexes might be organised in order to control electron transport pathways. Knowing the answers to these questions could give us powerful new tools for controlling biological energy conversion, allowing the engineering or organisms suitable for more efficient biofuel production, for example. We are using a cyanobacterium (a kind of photosynthetic bacterium) as a model organism. In cyanobacteria both photosynthesis and respiration occur in a complex membrane system inside the cell called the thylakoid membranes. Our initial approach to understanding the control of electron transport routes in cyanobacteria has used techniques in which we generate mutants in which particular electron transport proteins are fused to a fluorescent protein. We can then use a fluorescence microscope to see the distribution of electron transport proteins in intact membranes in living cells. A disadvantage of fluorescence microscopy is that it has relatively low resolution, i.e. we cannot see the organisation of electron transport components at molecular scales. However, the technique has allowed us to observe that the distribution of electron transport complexes is very heterogeneous on the scales that we can observe. Under some conditions particular electron transport components are concentrated into distinct patches in the membrane. We can show that the distribution of complexes into patches, or otherwise, is under physiological control, and we can relate this to the physiological control of electron transport. Our aims in this project are to understand better how the membrane patches are formed, what they contain, how they are organised and what effects they have on electron transport. Initially we will use an extension of our current fluorescent labelling techniques to get a more complete picture of the composition of the patches in living cells. This will be combined with studies to test hypotheses for the ways in which the distribution of complexes in the membrane are under physiological control. We will then use biochemical techniques to isolate the membrane patches and determine their full composition, combined with the use of electron microscopy to get higher-resolution information on the organisation of protein complexes in the membrane. At the end of the study we expect to understand much better how electron transport in the membrane is controlled. We expect our studies on a cyanobacterium to act as an exemplar for studies of biological electron transport at the membrane scale in other organisms, and to provide new ideas for the control of the organisation and function of biological membranes in general.