Cryo-EM structure of a membrane-embedded glutamate receptor

描述（由申请人提供）：结构生物学长期追求的目标是确定膜蛋白在其天然膜环境中的结构，以及由底物分子的跨膜梯度和膜电位定义的各种功能状态。这种梯度的应用，更不用说在膜环境中结晶，在x射线结构测定的情况下是不可能的。梯度的应用在电子晶体学的二维晶体分析中仍然是遥不可及的。提出的工作涉及进一步发展一种通用方法，用于单颗粒低温电镜结构测定重组成脂质体的膜蛋白。这种方法被称为随机球约束（RSC）单粒子重建。在我们提出的工作中，我们将努力克服几个剩余的技术障碍，然后将这种生物化学，EM成像和图像处理方法的结合应用于将GluA2谷氨酸受体重组为脂质膜的纳米分辨率结构。

DESCRIPTION (provided by applicant): A long-pursued goal of structural biology has been the determination of the structure of membrane proteins in their native membrane environment, and in their various functional states as defined by transmembrane gradients of substrate molecules and membrane potentials. Application of such gradients, let alone crystallization in a membrane environment, has not been possible in the case of X-ray structure determination. The application of gradients has remained out of reach as well for the analysis of 2D crystals by electron crystallography. The proposed work concerns the further development of a general method for single-particle cryo-EM structure determination of membrane proteins reconstituted into liposomes. The method is called random spherically- constrained (RSC) single-particle reconstruction. In the proposed work we will work to surmount several remaining technical hurdles, and then apply this combination of biochemistry, EM imaging and image processing methods toward a nanometer- resolution structure of the GluA2 glutamate receptor as reconstituted into a lipid membrane.