CHOLECYSTOKININ A RECEPTOR

从一系列的突变和嵌合体研究中 缩胆囊素(CCK)是一种肽类激素，与 神经递质具有广泛的生理作用，一直以来 定位于G蛋白偶联受体的胞外区。 为了获得这一生物分子相互作用的结构基础 激素和受体，我们合成了由47个氨基酸组成的多肽 受体N-末端结构域的序列(约7 第一个跨膜螺旋的氨基酸残基，作为 膜锚)。我们的目标是从结构上描述这一点 存在和不存在天然多肽激素的区域 (截断研究表明，C-末端有七个残基 的天然激素足以结合和药理作用 CCK的行动)。受体结构域是用光谱分析的 在DPC胶束存在下模拟膜的研究 环境。受体结构域的质子谱非常 由于大量的疏水残留物而拥挤。.的存在 缓慢滚动的胶束导致了显著的加宽 需要使用高场仪器的质子信号。 光谱使我们能够识别出一个独特的折叠 N-末端，形成推测的CCK激素结合袋。 该折叠由两亲性的β-折叠组成，由一个 两个半胱氨酸残基之间的二硫键，允许原子 深入了解已对 G蛋白偶联受体。所有这些都是通过 隔离的N端结构域的结构。

From a series of mutagenesis and chimeric investigations the binding site of cholecystokinin (CCK), a peptide hormone and neurotransmitter with a wide spectrum of physiologic actions, has been located to the extracellular domain of the G-protein coupled receptor. To obtain a structural basis for the biomolecular interaction of this hormone and receptor, we have synthesized the 47-amino acid peptide sequence of the N-terminal domain of the receptor (approximately 7 amino acid residues of the first transmembrane helix, to serve as a membrane anchor). Our goal is to structurally characterize this region both in the presence and absence of the natural peptide hormone (truncation studies have indicated that the C-terminal seven residues of the native hormone are sufficient for binding and pharmacological action of CCK). The receptor domain was spectroscopically investigated in the presence of DPC micelles to mimic the membrane environment. The proton spectrum of the receptor domain was very crowded due to an abundance of hydrophobic residues. The presence of the slowly tumbling micelles resulted in a significant broadening of the proton signals which required the use of a high field instrument. The spectra have allowed us to identify a unique fold of the N-terminus that forms the putative binding pocket for the CCK hormone. This fold consisting of an amphipathic beta-sheet, stabilized by a disulfide bridge between two Cys residues, has allowed for atomic insight into the modifications which have been carried out on the G-protein coupled receptor. All of this has been obtained with the structure of the isolated N-terminal domain.