A structural context for the mechanism of Uncoupling protein-1

我们从糖、脂肪和其他食物成分的分解中获得的大部分能量是由线粒体利用的，线粒体是我们细胞的“发电厂”。然而，能量转换的过程并不是100%有效的。在哺乳动物的特殊“棕色脂肪”细胞中，这一过程被主动“短路”，以燃烧热量。这些细胞使用一种叫做解偶联蛋白1 （UCP1）的特殊蛋白质来完成这一过程。利用UCP1产热对包括牲畜在内的许多哺乳动物，尤其是新生儿，在寒冷的温度下生存非常重要。虽然它对成年人也有好处。活跃的棕色脂肪往往出现在较瘦的人身上，他们不太可能患上与年龄有关的肥胖，这与消耗多余卡路里和改善健康状况的组织是一致的。棕色脂肪中的UCP1活性有助于清除血液中的葡萄糖和脂肪，这有助于对抗糖尿病。目前，肥胖和相关的不良健康状况正日益成为一个重大的国家和国际问题，迫切需要更好地了解新陈代谢。现在，全世界都在进行重要的科学研究，以鼓励棕色脂肪的发展，作为增加卡路里消耗以促进健康的可行手段。然而，棕色脂肪的一个重要特征是UCP1在细胞中并不是天生活跃的，必须特别“打开”才能燃烧卡路里以获得热量。这通常是通过调节分子的作用来发生的，这些调节分子激活UCP1，以响应生理刺激，如低温。在没有生理刺激的情况下，人工激活UCP1的治疗策略有可能大大增加组织燃烧卡路里的能力。然而，目前我们还不知道UCP1是如何工作的，也不知道调节分子是如何与蛋白质相互作用来启动它的。UCP1是一种膜蛋白，由于其不溶性和分离时的不稳定性，通常难以研究。然而，最近，我们对与UCP1相关的膜蛋白如何工作的理解取得了一些关键进展，这为确定UCP1的分子机制提供了新的机会。我的研究提供了纯化和评估UCP1的新方法，揭示了蛋白质的基本功能单元和稳定它的相关脂质分子。这里概述的工作旨在利用这些进展并阐明UCP1的分子性质。这种蛋白质将从天然来源（新生羔羊）和经过工程改造的酵母中纯化出来。关于调节分子如何与分离的蛋白质相互作用的细节将使用生物物理方法进行研究，该方法监测结合事件释放的热量或发生的蛋白质稳定性的相关变化。UCP1也将被整合到人造“脂质体”膜中，这样就可以评估调节因子如何影响其活性的细节。遗传方法将用于改变蛋白质的关键部分，以帮助确定激活分子结合的位置，以及蛋白质其他显著结构特征的功能。此外，将进行UCP1结晶实验，并使用x射线晶体学方法获得蛋白质外观的详细图片。这些研究将为UCP1使用的分子过程提供有价值的见解，这将推进我们对膜蛋白如何工作的基本理解，以及细胞中能量代谢的控制方式。揭示的细节可能为识别参与UCP1激活的其他细胞成分提供线索，并且对于激活针对肥胖和代谢疾病的能量消耗途径的治疗策略的合理化也将是无价的。

Most of the energy that we gain from the breakdown of sugars, fats and other food components is harnessed by mitochondria, the 'power plants' of our cells. However, the process of energy conversion is not 100% efficient. In specialised 'brown fat' cells of mammals, the process is pro-actively 'short circuited' to burn off calories as heat. These cells do this using a particular protein called uncoupling protein 1 (UCP1). Heat production using UCP1 is important for many mammals including livestock to survive cold temperatures, especially newborns. Though it is also beneficial for adult humans. Active brown fat tends to occur in leaner people who are less likely to develop age-related obesity, consistent with the tissue expending excess calories and improving health. UCP1 activity in brown fat helps remove glucose and fat from the blood, which can help combat diabetes. At present, obesity and related poor health conditions are growing as a major national and international problem for which a better understanding of metabolism is urgently needed.There is now significant scientific research worldwide into ways to encourage the development of brown fat as a viable means to increase calorie-expenditure to deliver better health. However, an important feature of brown fat is that UCP1 is not inherently active in the cells and must be specifically 'switched on' to burn calories for heat. This normally occurs through the action of regulator molecules that activate UCP1 in response to physiological stimuli such as cold temperatures. Therapeutic strategies to artificially activate UCP1 in the absence of physiological stimuli have the potential to greatly increase the capacity of the tissue to burn calories. Yet at present we do not know how UCP1 works nor how the regulator molecules interact with the protein to turn it on.UCP1 is a membrane protein, which are generally difficult to study due to their insoluble nature and instability when isolated. Recently, however, there have been some key advances in our understanding of how a membrane protein related to UCP1 works, which provides new opportunities to determine the molecular mechanism of UCP1. My investigations have provided new methods to purify and assess UCP1, which have revealed the basic functional unit of the protein and the associated lipid molecules that stabilise it. The work outlined here aims to capitalize on these advances and clarify the molecular nature of UCP1. The protein will be purified from natural sources (newborn lambs) as well as yeast that have been engineered to make it. Details on how regulator molecules interact with the isolated protein will be investigated using biophysical methods that monitor either the heat released from the binding event or the associated changes in the protein's stability that occurs. UCP1 will also be incorporated into artificial 'liposome' membranes, so that details on how regulators influence its activity can be assessed. Genetic methods will be used to alter key parts of the protein to help determine where activating molecules binds, as well as the function of the protein's other notable structural features. Additionally, experiments to crystallise UCP1 and use x-ray crystallography methods to gain a detailed picture of what the protein looks like will be carried out.These studies will provide valuable insight into the molecular process used by UCP1, which will advance our fundamental understanding of how membrane proteins work but also the way in which energy metabolism is controlled in cells. The details revealed may provide clues to identify other cellular components involved in UCP1's activation, and will also be invaluable to rationalise therapeutic strategies to activate energy expenditure pathways for targeting obesity and metabolic disease.