CONFORMATIONAL DYNAMICS IN ION CHANNEL SELECTIVITY AND GATING

 描述（由申请方提供）：离子通道控制细胞中的静息膜电位和动作电位。因此，离子通道的不适当的生理功能导致多种疾病状态，包括心脏和神经功能障碍、癫痫、哮喘、听力损失等。适当的离子通道功能需要适当的离子选择性和通道门控以控制穿过生物膜的离子通量。然而，我们对离子通道的这些基本性质的理解仍然是不完整的，因为它们来自于结构和动力学的复杂相互作用。我们将使用溶液核磁共振实验测试的作用，蛋白质动力学的离子选择性和门控的钠钾通道从蜡状芽孢杆菌溶解在各向同性bicelles。NaK是一个理想的模型系统，因为小的突变可以改变其选择性;它的结构与真核生物的钾类似 通道孔结构域，特别是生物医学上重要的环核苷酸门控和HERG钾通道;并且它高度稳定，易于进行结构和生物物理表征。NMR提供了一种独特的方法，同时和定量探测离子通道的结构和动力学与特定位点的分辨率。NMR的能力，以测量动态的不同的时间尺度和检测瞬态和低人口状态，使我们能够实验评估蛋白质动力学的作用，从离子选择性通道门控的过程。在这个建议中，我们将使用核磁共振研究钠钾测试三个目标：选择性产生的离子结合的骨干动力学的合作效应？N-末端M0螺旋如何调节NaK门控？什么样的结构和动力学特征允许钾通道中选择性过滤器和内门之间的变构通讯？

 DESCRIPTION (provided by applicant): Ion channels control both the resting membrane potential and the action potential in cells. As a result, improper physiological function of ion channels leads to a wide variety of disease states, including cardiac and neurological dysfunction, epilepsy, asthma, hearing loss, and others. Proper ion channel function requires appropriate ion selectivity and channel gating to control ion flux across biological membranes. However, our understanding of these fundamental properties of ion channels is still incomplete because they arise from a complex interplay of structure and dynamics. We will use solution NMR to experimentally test the role of protein dynamics in ion selectivity and gating of the NaK channel from Bacillus cereus solubilized in isotropic bicelles. NaK is an ideal model system because small mutations can shift its selectivity; its structure is similar to eukaryotic potassium channel pore domains, particularly biomedically important cyclic nucleotide gated and HERG potassium channels; and it is highly stable and amenable to structural and biophysical characterization. NMR provides a unique method to simultaneously and quantitatively probe ion channel structure and dynamics with site-specific resolution. The ability of NMR to measure dynamics on diverse timescales and detect transient and lowly populated states allows us to experimentally evaluate the role of protein dynamics on processes ranging from ion selectivity to channel gating. In this proposal we will use NMR studies of NaK to test three aims: Does selectivity arise from cooperative effects of ion binding on backbone dynamics? How does the N-terminal M0 helix regulate NaK gating? What structural and dynamic features allow allosteric communication between the selectivity filter and inner gate in potassium channels?