Structure and Function of Neurotransmitter Transporters

描述（由申请人提供）：中性和阴离子氨基酸的转运蛋白在人体生理中起着关键作用，在肾脏和大脑等多种器官中都很活跃。在中枢神经系统中，谷氨酸介导大多数快速兴奋性信号，这是一种神经元间的通信形式，对神经系统的发育和维持至关重要。谷氨酸介导的信号传导的一个基本组成部分是神经细胞和其他细胞（如神经胶质细胞）中依赖钠的高亲和力谷氨酸转运体在兴奋性刺激后将谷氨酸从突触间隙中移除。目前，还没有关于谷氨酸转运体的原子分辨率结构信息，这极大地阻碍了我们对其结构和作用机制的理解。在本应用中，我建议使用x射线晶体学来确定与真核谷氨酸转运体具有显著序列同一性的细菌蛋白的结构。此外，我计划确定细菌同源物的功能行为，并测试基于结构的转运蛋白功能机制。此外，通过使用细菌蛋白的晶体结构作为指导，我将创建选定真核转运蛋白的同源模型，并与先前确定的结构和功能信息一起，这将把真核转运蛋白的结构和功能关系置于原子分辨率的三维环境中。综上所述，本研究将进一步加深我们对真核生物和原核生物谷氨酸转运体的认识，并且由于它们与二羧酸转运体和中性氨基酸转运体有关，我们对这些二级转运体的认识也将增加。最后，由于谷氨酸能信号在人类神经系统中普遍存在，细菌转运体的结构，以及真核转运体的同源模型，应该有助于设计可能具有治疗潜力的新分子。

DESCRIPTION (provided by applicant): Transporters for neutral and anionic amino acids play keys roles in human physiology and are active in organs as diverse as the kidney and brain. In the central nervous system, glutamate mediates the majority of fast excitatory signaling, a form of neuron-neuron communication that is essential to the development and maintenance of the nervous system. A fundamental component of glutamate-mediated signaling is the removal of glutamate from the synaptic cleft, following an excitatory stimulus, by sodium-dependent, high affinity glutamate transporters in neurons and other cells, such as glial cells. At the present time, there is no atomic resolution structural information on a glutamate transporter, which greatly hampers our understanding of their architecture and mechanism of action. In this application I propose to determine the structure of a bacterial protein that has significant sequence identity to the eukaryotic glutamate transporters, using x-ray crystallography. Furthermore, I plan to determine the functional behavior of the bacterial homolog, and to test structure-based mechanisms of transporter function. In addition, by using the crystal structure(s) of the bacterial protein as a guide, I will create a homology model of selected eukaryotic transporters and, together with previously determined structure and function information, this will place structure and function relationships of the eukaryotic transporters in an atomic-resolution, three-dimensional context. Taken together, the proposed research will substantially further our understanding of both eukaryotic and prokaryotic glutamate transporters, and, because they are related to transporters of dicarboxylic acids and of neutral amino acids, our knowledge of these secondary transporters will be increased as well. Lastly, because glutamatergic signaling is pervasive in the human nervous system, the structure of the bacterial transporter, along with the homology models of the eukaryotic transporters, should facilitate the design of new molecules that may have therapeutic potential.