Structure and Function of the Acetylcholine Receptor

我们研究的长期目标是了解神经递质受体在健康和疾病状态下的运作方式。我们选择运动突触上的乙酰胆碱受体（AChR）来研究这类蛋白质的基本功能：结合神经递质触发内在离子通道的打开。为了理解这一结合触发过程，我们建议(i)识别a和非a亚基中的残基，这些残基在结合到通道的打开和关闭状态时稳定ACh， （ii）确定AChR如何维持均匀的通道门控动力学，（iv）确定先天性肌弱综合征（CMS）突变的机制后果。（iv）确定在主要胞外结构域的推定二级结构的保守残基是否有助于AChR的激活；(v)确定AChR的主要胞外结构域的结构特征。该方法结合了位点定向突变和哺乳动物细胞的表达、单通道记录和动力学分析、配体结合和蛋白质生物化学的测量。这些研究的完成将促进我们对运动终板突触传递和药物作用的理解，促进神经肌肉疾病（如CMS）的治疗，而总体研究结果将为神经递质受体超家族其他成员的结构功能关系提供见解。

The long-term aim of our research is to understand how receptors for neurotransmitters operate in health and disease states. We have chosen the acetylcholine receptor (AChR) at the motor synapse to examine the essential function of this class of proteins: binding of neurotransmitter triggering opening of an intrinsic ion channel. Toward understanding this binding-triggering process, we propose to (i) identify residues in a and non-a subunits that stabilize ACh when bound to open and closed states of the channel, (ii) determine how the AChR maintains uniform channel gating kinetics, (iv) determine mechanistic consequences of mutations underlying congenital myasthenic syndromes (CMS), (iv) determine whether conserved residues bounding putative secondary structures in the major extracellular domain contribute to AChR activation and (v) determine structural features of the major extracellular domain of the AChR. The approach combines site-directed mutagenesis and expression in mammalian cells, single channel recording and kinetic analysis, measurements of ligand binding and protein biochemistry. Completion of the proposed studies will advance our understanding of synaptic transmission and drug action at motor endplates, facilitate treatment of neuromuscular disorders such as CMS, while the general findings will provide insight into structure function relationships for other members of the neurotransmitter receptor superfamily.